Histology Laboratory Manual: Olgga A. Hara MS

Histology
Laboratory Manual
Olgga A. Hara MS
Assistant Professor
Biological Sciences Department
De La Salle University - Dasmariñas
S.Y. 2013 – 2014
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Contents
Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . iii
The use and care of the microscope . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . iv
Laboratory Activities
1. The Cell: Structure and Function . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
2. Epithelial Tissue. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
3a. Loose and Dense Connective Tissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
3b. Specialized Connective Tissues: Cartilage and Bones . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
4. Muscular and Nervous Tissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27
5. Blood and Lymphatic Tissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36
6. Integumentary system . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47
7a. Digestive System (Part 1) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ... . . . . . . . . . . . . . . . . . . 51
7b. Digestive System (Part 1) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ... . . . . . . . . . . . . . . . . . . 55
8. Central Nervous System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
Appendix: Histotechnique. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
Preface
Histology is a highly visual science. It is about looking at structures that
needs a keen recognition of patterns in tissues if the aim is to understand the
functioning human body. Normally, these patterns occur as organized cells and
their subcellular structures as well as the extracellular matrix that make up the
tissues in a healthy organ. Likewise, one’s knowledge and familiarity to normal
tissues is basic to the recognition and understanding of abnormal patterns seen
in pathologic tissues. Much can be learned about the condition of cells and
tissues by examining the details of their components, thus, structure is said to
be a useful hint to the function of an organ. However, a fuller knowledge of
tissue biology can be obtained when histology is associated with other
information such as tissue chemistry, physiology, immunology and other basic
sciences.
The Use and Care of the Microscope
I. How to focus a specimen under the microscope.
a. Place the prepared slide on the stage and secure the
slide in place with the clip or slide holder.
b. Manually position the specimen at the center by using
the mechanical stage controls.
c. Turn the revolving nosepiece so that the scanner (4X)
or LPO (10X) is in place and aligned with the stage
aperture.
d. Raise the stage by moving the coarse adjustment knob
(larger) as you look through the side.
e. Look through the ocular and as the illuminator is
turned on, adjust the light so that the field is bright
without hurting the eye. With the scanner or low
power objective you may have to reduce illumination
intensity. Figure 1. A Compound Light Microscope
f. Adjust the condenser by moving it as close to the stage aperture as you can get it. By adjusting it along with the iris diaphragm
through its lever, you will notice the light that comes up through the specimen change brightness.
g. Gradually lower the stage using the coarse adjustment knob until a sharp focus of the specimen is achieved.
h. Once the object is focused at LPO, you may need to magnify a part to see its details. Shift to High Power Objective by turning
the revolving nosepiece. Be sure you feel a click so that the objective lens is properly aligned to the condenser.
i. Do not move the coarse adjustment knob this time, but use the fine adjustment knob (smaller) instead. If the microscope has a
parfocal quality, it means you can readily shift to the next higher power objective with minimal adjustment because the
specimen remains in focus or close to focused.
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Note: As you move up to a higher power objective, you may need to re-focus and re-center the specimen.
j. When finer details must be resolved, you may shift to the Oil Immersion Objective (OIO). Before positioning this objective in
place, drop a small amount of immersion oil on the specimen. The tip of the objective must touch the oil at this point and adjust
by gradually moving the fine adjustment knob.
k. Increase light intensity as necessary especially when using the HPO and OIO but be sure you view the object with proper
contrast. Stained specimen can be focused with ease but in unstained specimen, you may shift to dark-field mode or simply
lessen the light intensity by having the iris diaphragm partially closed down.
II. Care of the microscope

a. Be careful with every manipulation so that the microscope will maintain its good quality.
b. When transferring a microscope, hold it firmly with one hand on the arm and the other by the stand. Never grab it by the
eyepiece holder or carry with one hand while swinging it.
c. Focus smoothly; don't try to speed through the focusing process or force anything.
d. The stage and lenses must be clean before putting away the microscope.
e. To clean the lenses, only a lens tissue or a cotton swab soaked in appropriate lens cleaner or distilled water can be used. Organic
solvents may separate or damage the lens elements or coatings. NEVER use a paper towel, a tissue paper, your lab suit, or any
material.
f. Turn the illuminator off when you are done or whenever you don’t use it.
g. Hold the plug (not the cable) when unplugging the illuminator.
h. Wrap the cord neatly around the microscope when you return them to the stockroom.
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1. The Cell: Structure and Function
The invention of the microscope led to the discovery of the microscopic world including the cell, which is the basic structural and
functional unit of all living organisms. A single cell may constitute within it all of the machinery to become one functional organism, as in the
case of protozoans. These single-celled organisms are capable of performing specialized functions such as movement, procurement of
food, responding to environmental stimuli, reproduction and waste elimination. However, a cell could also be one in a group of cells that
undergo development and differentiation to become specialized for a particular function. Hence, these cells appear to have distinctive
features that are grouped together to compose “tissues”; the tissues that make up organs, and organs that make up organ systems, and
organ systems that make up the entire multicellular organism. Therefore, whenever cellular components are destroyed by any mechanical
or structural means, cellular function is likewise destroyed as it is the fundamental structural and functional unit.
All animal cells share a common basic structure: the outer covering plasma membrane/cell membrane, the nucleus that contain the
genetic material in the form of chromosomes, and the cytoplasm that bears the small structural elements termed organelles. All these
cellular components provide the framework for cellular activities. One learns about these structures by observation of cytological
preparations or tissue details under the microscope.
Tissues are prepared as thin sections that are mounted on glass slides and stained with appropriate dyes because tissue elements
are virtually colorless under the light microscope. A combination of hematoxylin and eosin (H and E) stains are commonly used for basic
histologic studies. Hematoxylin is a basic stain and the cellular structures that it stains are termed basophillic, hence the color imparted is
purple. Contrary to eosin which is an acid stain, wherein the structures that it stains are termed acidohillic or eosinophillic, hence the color
imparted is pink. In the H and E technique, cell nuclei and other cytoplasmic components stain purple while the cytoplasm generally stain
pink.
A keen observation of the normal appearance of cells in the body through histological preparations is very important if abnormal
cells due to certain diseases are to be recognized. Aside from gaining insight into the structure and function of organs and systems,
studying cells is essential in the understanding of normal as well as the abnormal biochemical and physiological processes in the body.
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Activity 1. The Cell: Structure and Function
Objectives: At the end of this activity the student is expected to:
1. Identify different kinds of cells and recognize subcellular structures as seen under the microscope.
2. Describe how hematoxylin and eosin (H & E) stains make the various components of cells and tissues visible.
3. Relate the subcellular components with the primary function of the cells in a tissue.
4. Recognize abnormalities or changes in a cell as a result of pathologic factors.
5. Realize the importance of maintaining healthy cells for an efficiently functioning body.
Materials:
Microscope, immersion oil, lens paper, cotton, xylene
Prepared slides: frog spinal cord section, human squamous epithelium, liver section: normal and pathologic sections
Procedure:
A. Frog Spinal Cord, c.s.
1. Focus on the gray matter area in the cross section of a frog spinal cord. Under the scanner objective, this structure appears as a
butterfly-shaped region at the center.
2. Magnify the gray matter area using LPO to locate nerve cell bodies of motor neurons (stellate-shaped nerve cells) which are
scattered. Using the HPO may be necessary to view an entire cell body.
3. Examine a nerve cell body and identify the following structures:
Cell membrane – the lining that envelopes the cell. It is quite indistinct in H and E stained preparation.
Cytoplasm – the matrix within, that contains the organelles, inclusions and cytoskeleton, which are all kept in place. Take note of
the dominant characteristic staining reaction with H and E technique.
Nucleus - a relatively large, distinct spherical structure that is centrally located inside the cell body.
Nucleolus – prominent densely staining basophilic structure within the nucleus.
Cytoplasmic extensions - nerve cell processes that typically come out from the cell body as extension.
4. Draw a neuron cell body and label its parts in the worksheet (A).
B. Liver section
1. Examine the liver cells (hepatocytes) using the OIO of your microscope. The hepatocytes are seen as polyhedral cells that are
arranged in cord and plates radiating from the central canal in a spoke-like manner.
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2. Look for the cells with less-dense nuclear materials and determine the following structures:
Nucleus – spherical, centrally located body within the cell.
Nuclear membrane - separates the nucleus and the nucleolus from the rest of the contents of the cell.
Nucleolus - a dense spherical structure within the nucleus of a cell.
Nuclear sap/nucleoplasm – the matrix within the nucleus.
Chromatin materials – dense staining bodies that are attached to the nuclear membrane inside the nucleus.
Cytoplasm - the matrix outside the nucleus that contains the organelles, cell inclusions and cytoskeleton.
Cell membrane – the lining that envelopes the cell.
3. Draw a hepatocyte and label its parts in the worksheet (B).
4. Take note of the cytoplasmic granules and other components as seen with the light microscope.
C. Human squamous epithelium (inner-cheek lining cells)

1. Examine a smear of inner cheek lining epithelium stained with hematoxylin and eosin. Identify the following structures in the cells
using the HPO.
Nucleus - basophilic due to nucleic acids within it.
Nuclear envelope - basophilic due to adsorbed DNA.
Nucleolus - basophilic due to RNA.
Chromatin - basophilic due to DNA.
Cytoplasm – acidophilic due to the stain’s affinity to the basic components within it.
2. Draw a cheek lining cell and label its parts in the worksheet (C).
3. Describe each subcellular structure based on electron micrographed image of a cell and give their function. Complete the table in
the worksheet (D).
D. Pathologic tissue sections

1. Examine sections from normal and pathologic liver.
2. Differentiate the features in the cells of normal and pathologic liver section based on the following structures:
a. nuclear membrane - if distinct or indistinct.
b. nucleolus – if intact or dissociated.
c. nuclear sap/ nucleoplasm – based on consistency: if concentrated (darkstaining) due to condensed chromatins or fragmented
or faded in appearance
d. Cytoplasm – if abundantly granular in content or vesiculated (with large vacuoles) with decreased staining in appearance.
e. Cell size – diameter: if shrunken or swollen or regular size.
3. Complete the table in the worksheet (E).
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WORKSHEET NO.1 Name ______________________
Course/Year & Section _________
Group No. ____
THE CELL: STRUCTURE AND FUNCTION Date submitted _______________

Professor ___________________
Rating _____________________
A. Draw and place arrows that correspond to the structures of a neuron cell body in the gray matter of frog spinal cord section.
Cell membrane Cytoplasm
Nucleus Cytoplasmic extension
Nucleolus
Neuron cell body

Mag. _________
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B. Draw and place arrows that correspond to the structures of a inner-cheek lining cells in the human squamous epithelium.
Cell membrane Cytoplasm
Nucleus Chromatin materials
Nucleolus
Squamous cell
Mag._________
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C. Draw and place arrows that correspond to the structures of a liver cell or hepatocyte.
Nucleus Chromatin materials
Nucleolus Cytoplasm
Nucleoplasm Cell membrane
Hepatocyte
Mag. ______
a. What organelles are most abundant in the cytoplasm of hepatocytes? What is the significance of these organelles in
the primary function of the liver?
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D. Give the special characteristics and basic functions of the following cytoplasmic organelles.
STRUCTURE SPECIAL CHARACTERISTICS FUNCTION
a. Golgi apparatus
b. Lysosomes
c. Mitochondria
d. Ribosomes
e. Nucleus
f. Smooth endoplasmic
reticulum
g. Rough endoplasmic
reticulum
h. Centrosomes
i. Microvilli
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E. Describe the cellular structures as seen in the hepatocytes of a liver section in the normal state and pathologic state.
DESCRIPTIONS
STRUCTURES
NORMAL PATHOLOGIC
a. nuclear membrane
b. nucleolus
c. nuclear sap/ nucleoplasm
d. Cytoplasm
e. Cell size
CONCLUSION
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2. Epithelial Tissues
The developing vertebrate embryo reveals the through microscopic examination. Therefore, epithelial
formation of germ layers as the cells rapidly divide. From tissues are primarily classified as simple epithelia whenever
each of these germ layers, four types of tissue develop, the layer is just one-cell thick; or as stratified epithelia
persist to maturity and constitute the various organs in the whenever there are two or more layers of cells. The cell
body. These are called epithelial, connective, muscular and shape on the other hand, gives the secondary classification
nervous tissues. as being squamous type (flattened cells), cuboidal type
Characterized by cells that are closely apposed to (cube-shaped cells) or columnar type (tall cells).
each other, the epithelial tissue is devoid of any intercellular The epithelial tissues not only serve as lining or
substance and is completely avascular. They cover and line covering structures in the body. They are composed of cells
all the body surfaces, cavities and tubes. Those that line the that are active in performing essential roles in the body such
large internal cavities are called mesothelium and those that as absorption of digested food, secretion of hormones and
line the internal surfaces of blood and lymph vessels are other substances, sensation, temperature regulation, and for
called endothelium. The underlying connective tissue is protection. The secretory function entails another category
where the epithelial cells depend for anchorage and of epithelial tissues called glands. Glands are
nourishment through diffusion of substances. The epithelium developmentally derived from the epithelia, which form a
and its underlying connective tissue are separated by a down growth into the underlying connective tissue. They are
structure called the basement membrane. composed of cells or aggregation of cells, which have the
In recognizing epithelial tissues, one must be able to ability to secrete certain substances. Glands are highly
discern the number of cell layers as well as the cell shape functional epithelia due to their secretory activity.
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Activity 2 – Epithelial Tissues
1. Recognize the different tissue types in a set prepared slides of different organs of the body.
2. Associate the characteristics of tissue types to their functions in the organs where they are found.
3. Identify some peculiar structures unique in each type of tissue.
Materials:
Prepared slides: Kidney section, stomach, human brown skin, esophagus, salivary gland, trachea, urinary bladder.
Procedure:
A. EPITHELIAL TISSUES
Recognize the type of epithelium in a tissue according to:
1. Layers – simple, stratified, pseudostratified
2. Shape of cells – squamous, cuboidal, columnar, transitional
3. Presence of any surface modification/ specialization – cilia, microvilli (brush borders), stereo cilia, keratin
4. Function - Covering epithelia, Lining epithelia, Absorptive epithelia, Secretory or Glandular epithelia (exocrine and endocrine
glands)
In giving the complete name of an epithelial type, follow the order given below:
Identify LAYER/S + Identify SHAPE of cells (topmost cells for stratified) + Identify if there is a SURFACE MODIFICATION present
Example: Pseudostratified columnar ciliated epithelium
1. Examine the following slides and locate the given structures where the epithelial tissue is found.
1.1. SIMPLE EPITHELIUM
a. Kidney section: Bowman’s capsule
Scan through the cortical parenchyma (peripheral region) of the kidney section and you will find several dense, rounded structures
called renal corpuscles surrounded by convoluted tubules that were transversely sectioned. The dense, rounded structures
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are the glomeruli within the Bowman’s capsules. A higher magnification is required to determine the type of epithelium
present in the lining of the capsule as well as the glomerulus within it.
Look for the following structures:
Lining of the Bowman’s (glomerular) capsule – composed of flattened cells surrounding the glomerular capillaries.
Glomerulus – a globular network of anastomosing capillaries, which invaginates Bowman’s capsule.
Draw a portion of the lining tissue of the Bowman’s capsule, identify the type of epithelium and its function.
b. Kidney section: Renal tubules of the cortical parenchyma.

In the same cortex region of the kidney, surrounding the renal corpuscles are convoluted tubules. Focus on these tubules under the
HPO and describe the cells lining them.
Draw the epithelium of the renal tubules in the renal cortex, identify the type of epithelium and its function.
c. Gall bladder: Fingerlike folds of the mucosa

Focus on the finger-like folds of the mucosa under the HPO. Take note of the layer of the nuclei and the cytoplasm of the apical
lining cells.
Columnar cells – a layer of tall epithelial cells with elongated nuclei, which exhibit basal polarity. The cytoplasm seems to be
acidophilic. The luminal plasma membrane is lined with brush borders (microvilli).
Basement membrane – indistinct underlying structure at the basal region of the epithelial cells.
Draw the mucosa of this structure, identify the type of epithelium and give its function.
1.2. STRATIFIED OR COMPLEX TYPE

a. Human brown skin: Epidermal layer
Focus on the epidermal layer of the skin under the LPO. Take note of the layer of flattened cells. The cells near the base have
rounded nuclei. Dead cells lose their nuclei.
Draw the epidermal layer and identify the type of epithelium and give its function.
b. Parotid gland: Larger excretory duct/ interlobular ducts

The gland parenchyma is divided into lobes and lobules by septa called interlobular connective tissue septa. Focus on this area and
look for interlobular excretory duct, which are larger in diameter than the other similar structures you can find. Examine the
layers of the lining cells.
Draw the larger excretory duct, identify the type of epithelium and give its function.
c. Parotid gland: Main excretory ducts/ interlobar ducts
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Using LPO, locate the interlobar connective tissue. This is the area that separates the main lobes of the secretory areas of the gland.
The excretory ducts from the lobules join to form the main excretory duct. Look for the main excretory duct and examine
the layers of the lining cells using HPO.
Draw the larger excretory duct, identify the type of epithelium and give its function.
d. Trachea: Inner Lining

Using the HPO, focus on the inner lining of the trachea. Notice the arrangement of the cell nuclei and the cytoplasm of the cells.
Columnar cells – a layer of cells which attach to a common basement membrane but appear to form several layers due to
the nuclei of cells located at different heights within the epithelium.
Cilia – hair like structures covering the epithelial cells’ apical part except those of the goblet cells.
Basement membrane – connective tissue lining which is clearly visible, separating the pseudostratified epithelium from the
underlying connective tissue.
Draw the tracheal inner lining, identify the type of epithelium and give its function.
e. Urinary bladder: Lining of the mucosa

Focus on the lining of the mucosa of this organ under LPO. A higher magnification may be used to see the shape of the cells and
their arrangement in layers.
Look for the different layers of cells in the epithelia and other structures:
Surface cells – usually cuboidal cells that bulge outward; convex, dome-shaped luminal surface and frequently
binucleate (with two nuclei).
Intermediate cells – polyhedral; several layers in thickness.
Basal cells – deeper layers of rounded or oval cells
Basement membrane - connective tissue lining which is clearly visible, separating the epithelium from the underlying
connective tissue.
Lamina propia - loose connective tissue found immediately beneath the basement membrane.
Draw the lining of mucosa, identify the type of epithelium and give its function.
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WORKSHEET NO.2
Name ______________________
Group No. ____
Date submitted _______________
Epithelial Tissues Professor ___________________
Rating _____________________
A. EPITHELIAL TISSUES
SIMPLE EPITHELIUM
PSEUDOSTRATIFIED
SIMPLE SQUAMOUS SIMPLE CUBOIDAL SIMPLE COLUMNAR
COLUMNAR (CILIATED)
Tissue structure / slide Tissue structure / slide Tissue structure / slide Tissue structure / slide
Function Function Function Function
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STRATIFIED EPITHELIUM
STRATIFIED SQUAMOUS STRATIFIED CUBOIDAL STRATIFIED COLUMNAR TRANSITIONAL
Tissue structure / slide Tissue structure / slide Tissue structure / slide Tissue structure / slide
Function Function Function Function
CONCLUSION
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3. Connective Tissues
The connective tissues are a class of tissue due to their embryonic tissue origin, which is the mesoderm. Upon
development, these tissues have three basic components, which vary in amount and composition. These components are the
following: (1) character cells, (2) tissue fibers and the (3) ground substance. They provide structural and metabolic support for
other tissues and organs of the body. Bone and cartilage are best for support, protection of vital organs and serve as site for the
formation of blood cells. Unlike the epithelial tissues, there are blood vessels in the connective tissues through which the
exchange of nutrients, metabolites and waste materials between the tissues and the circulatory system is facilitated. Another
special function is the protection from foreign matters that may cause harm to the tissues by initiating inflammatory reactions
and through cellular and humoral defense mechanism. Through the blood vessels, the immune cells may enter the connective
tissues and migrate to the site of infection that enable them to perform their protective function.
CONNECTIVE TISSUES (CT)

CT Components:
1. CT cells/ specialized support cells – fibroblasts, blood cells (red blood cells, white blood cells, plasma cells),
macrophages, mast cells
2. Extracellular matrix / ground substance – glycosaminoglygans, proteoglycans, water, salts, etc.
3. CT fibers – collagen, elastic, reticular fibers
CT Type according to density and pattern of fiber arrangement:
1. Loose CT – areolar, reticular
2. Dense CT – dense regular, dense irregular
3. Specialized CT
a. Cartilage – hyaline, elastic and fibrocartilage
b. Bone – Spongy bone, compact bone
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Activity 3a – Loose and Dense Connective Tissues
Types of Connective tissues
I. Loose connective tissue is characterized by its being cell-rich, soft and compliant tissue due to its abundant ground substance,
which fill the spaces and confer physical properties and control transport. It is also rich in blood vessels and nerves but
limited in fibers. This type of tissue may occur in some special variants:
Areolar connective tissue – consists of loosely arrange network of different type of fibers upon which many kinds of fixed
and wandering cells are suspended.
Reticular connective tissue – consists of an open meshwork of fine, reticular fibers that are formed by reticular cells or
reticulocytes. The open meshwork of fine fibers is particularly useful in tissues and organs in which diffusion and/or cell
movements are functionally important.
Adipose connective tissue – consists of large numbers of adipocytes or fat cells. Two types of adipose tissues can be
distinguished based on the color of the tissue and the number of lipid droplets found in the adipocytes:
Adipocytes of white, unilocular adipose tissue contain one large lipid droplet.
Adipocytes of brown, multilocular adipose tissue contain many lipid droplets.
II. Dense connective tissue is characterized by its being completely dominated by tissue fibers. The tissue can be further
categorized based on the spatial arrangement of the fibers in the tissue.
Dense regular connective tissue – if the fibers run parallel to each other.
Dense irregular connective tissue – if the fibers do not show a clear orientation within the tissue but instead form a densely
woven three-dimensional network.
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Materials:
Prepared slides: Human brown skin, Areolar tissue (mesentery spread), lymph node section, tendon
Procedure:
Examine the following prepared slides and recognize the connective tissue components and complete the table in the worksheet.
a. Mesentery spread
Look for the following structures, identify the CT type and its main components.
Collagen fiber – coarse, thick fibers that cross one another which usually stain pink or eosinophillic.
Elastic fiber – dark staining branching fibers that are more slender than collagen fibers.
Mast cell – large cells with coarse cytoplasm and small, round nucleus.
Fibroblast – found to be the most abundant type of cell in this tissue, the cell membrane of these cells are indistinct in H & E.
Only the large, ovoid nuclei are visible.
Lymphocytes – the nuclei are also round but smaller than fibroblasts and are more darkly stained.
b. Human brown skin: Hypodermal region

Fat/adipose cells – round to oval shaped, large cells that look like empty spaces due to the fats that are removed upon tissue
processing.
Nuclei of fat cells – small, dark staining structures that are eventually pressed against the cell membrane, giving more area
for fats in the cytoplasm.
Fibroblasts – cells with large, ovoid nuclei, prominent nucleoli and basophilic cytoplasm that are found near the adipose
cells.
c. Lymph node: Stroma

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Reticular fibers – unseen in H. & E. preparations but appear as dark staining fibers in special staining technique. This forms
complex branches of very fine fibers (collagen type 3) that crisscross the entire area of the lymph node especially
the cortex.
Reticular cells/ reticulocytes – somewhat stellate or star-shaped cells with processes that are attached to their adjacent cells.
The nucleus appear to be ovoid like that of fibroblast and the cytoplasm is light-staining.
Lymphoid cells – in the cellular parenchyma which are mostly lymphocytes. These are small cells with round, dark nucleus
that occupy mostly the entire cell, leaving little space for the cytoplasm.
d. Human brown skin: Dermal region

Elastic fibers – thin, intensely staining fibers that form a network that is dispersed irregularly.
Collagen fibers – thick, eosinophilic fibers that are randomly arranged and densely packed in this region.
Blood vessels – these are capillary loops situated right beneath the epidermis.
Character cells – mast cells that are found occasionally and fibroblasts that may be found in abundance.
e. Tendon section
Collagen fibers – dense aggregates of paralleled fibers that are separated by few areolar connective tissue having vessels
and nerves.
Tendon cells arranged in rows – these are fibrocytes, which show up only their prominent nuclei arranged in rows,
longitudinally oriented and found between the bundles of collagen fibers
Fibroblasts – these are mature fibrocytes which are stellate in shape, with ovoid nuclei and with cytoplasmic processes that
extend between and around the collagen bundles.
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WORKSHEET NO.3a Name ______________________
Loose and Dense Connective Tissues Group No. ____

Professor ___________________
Rating _____________________
A. CONNECTIVE TISSUE components

After examining the prepared slides, draw the components of connective tissues based on given descriptions in the
procedure.
COLLAGEN FIBERS ELASTIC FIBERS RETICULAR FIBERS FIBROBLASTS
LYMPHOCYTES PLASMA CELLS MACROPHAGES ADIPOCYTES/ FAT CELLS
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After examining the tissues, describe the type of CT and their main components by completing the table below.
Other sites with the same CT

Prepared slide CT Type Function
type
Holds intestines in place; to
a. Areolar supply blood vessels and
Mesentery spread nerves from central system to
the organ.
b. Human brown skin

Insulator of the body that
Fat, adipose layer in
reduce heat loss through the
the hypodermis of
skin; for energy storage.
skin
Reticular CT serves as fine

c. Lymph node section
supporting elements for the
(cortex region)
developing lymphocytes.
d. Dermis region of the

Gives flexibility and elasticity
skin
to the skin.
(reticular layer)
e. Tendon section A source of great tensile

strength
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Activity 3b – Specialized Connective Tissues
III. Specialised connective tissue is designated to cartilage and bones, the major skeletal components of the body.
Cartilage – consists of cells, especially chondrocytes and extracellular components. Unlike other connective tissues,
cartilage tissues are devoid of blood vessels or nerves. Cartilages are surrounded by perichondrium, which is a dense
connective tissue. Varying proportions of collagen and elastic fibers are embedded within the ground substance of this
tissue, which give rise to the three main types of cartilage:
Hyaline cartilage – occurs fused with bone or as discrete pieces, looking translucent to the unaided eye. This is due to the
abundance of collagen and its ground substance having similar refractive properties.
Fibrocartilage – consists of alternating layers of hyaline cartilage matrix and thick layers of dense collagen fibers.
Elastic cartilage – consists of numerous bundles of branching elastic fibers in the cartilage matrix, particularly in the
immediate vicinity of the chondrocytes.
Bone – is a rigid connective tissue with osteocytes or bone cells in a matrix of densely packed collagen fibrils infiltrated
with minerals. Like cartilage, it is surrounded by a dense connective tissue called periosteum. A marrow cavity is found
within this rigid tissue and the lining of this cavity is a thin layer of cell-rich connective tissue called endosteum. Both
periosteum and endosteum have bone cell-forming (osteogenic) potency. Unlike the cartilage, bone tissues are rich in
blood vessels, which run through long processes within the matrix called canaliculi. Bones are classified based on the size
of the spaces within it, and its trabecular or dense nature.
Spongy bone – consists primarily of slender, bony trabeculae branch and intersects to form a sponge-like network.
Compact bone – does not have any spaces or hollows in the bone matrix.
20
Materials:
Prepared slides: Trachea, epiglottis, fibrocartilage, spongy bone, compact (decalcified) bone.
Procedure:
Examine the following prepared slides and recognize the connective tissue components and complete the table in the
worksheet.
a. Trachea: Cartilage plate

Chondrocytes – cells having prominent nuclei which are rounded when situated at the center but appear to be
oval or flattened when at the periphery of the tissue. The cytoplasm appears to be vacuolated due to
removal of fats and glycogen droplets during tissue processing. These can be an individual cartilage cell,
sometimes found in pairs or groups within smooth-walled spaces.
Lacunae – these are the smooth-walled spaces that house the chrondrocytes.
Matrix – a special medium of ground substances through which cells and fibers are embedded. Most predominant
component in this cartilage tissue type with few collagen fibers that are not visible.
Isogenous groups of chondrocytes – cartilage cells that underwent mitotic division and the resulting cells are
grouped into small clusters.
Perichondrium – an outer layer of dense irregular connective tissue that ensheaths hyaline cartilage (except
articular cartilage in a synovial joint) and contain cells called chondroblasts.
Chondrogenic cells - stellate-shaped cells of fibroblast origin with spindle-shaped nucleus found interspersed
within the inner layer of the perichondrium. These cells give rise to chondroblasts.
b. Epiglottis
21
Perichondrium - an outer layer of dense irregular connective tissue that ensheaths the elastic cartilage.
Matrix with elastic fibers – similar with hyaline cartilage matrix although elastic fibers are readily visible and more
numerous. This cartilage is more pliable due to the abundance of elastic fibers that surround the
chrondrocytes.
Chondrocytes – similar cells as those of hyaline cartilage.
Lacunae - these are the smooth-walled spaces that house the chrondrocytes.
c. Intervertebral disc: Fibrocartilage

Collagenous fibers – eosinophilic fibers that are thick and run in parallel bundles. It is the predominant component
of this cartilage tissue type.
Matrix – very minimal in this cartilage tissue type.
Lacunae – the space where cartilage cells reside. Because of scarcity in matrix material, lacunae are usually
incomplete.
Rows of chondrocytes – cartilage cells within lacunae, somewhat arranged in regular rows (not in isogenous
groups) and found embedded in the matrix between collagen fibers. They are far less numerous and
muchmore widely separated than the other cartilage tissue types.
Nucleus of chondrocytes – round to ovally shaped basophilic structure.
Note: this cartilage type lacks an identifiable perichondrium.
d. Bone section: Spongy portion

Periosteum – a layer of tough/dense collagenous fibers that cover the compact bone shell. There are cellular
materials in its inner layer called osteogenic/osteoprogenitor cells which are spindle shaped.
Endosteum – a thin connective tissue layer with mixed cell types lining the marrow cavities and the spaces among
the trabeculae.
Trabeculae – sheets and plates of bone tissues found in the inner part of the section of bone surrounding the
marrow cavities.
Osteocytes – mature bone cells which are resident cells of the bone tissue that are entrapped within lacunae.
These cells have long processes that pass through the narrow canaliculi that radiate from the lacunae.
Lamellae – calcified matrix arranged in regular layers which surround the osteocytes in lacunae.
22
Bone cavity with myeloid tissues – the space within bones through which the network of trabeculae are inserted.
It is filled with marrow stuff which are the developing blood cells, adipocytes and fluids.
Adipose cells – Unilocular fat cells.
e. Bone, decalcified: Shaft of a long bone

Osteocytes - mature bone cells which are resident cells of the bone tissue.
Lacunae of osteocytes – the space produced when the matrix around osteocytes hardened.
Canaliculi – tiny hair-like channels that are branching. These radiate from every lacunae and the processes of
osteocytes pass through it so that one osteocyte can contact those adjacent ones.
Osteon – a unit which consists of concentric layers of calcified matrix (lamellae) where osteocytes are entrapped
around a central/Haversian canal. It is also referred to as the Haversian system.
Haversian canal – appear as a dark hole in cross section. It is a canal that is surrounded by concentric lamellae of
the compact bone. It conducts blood vessels, lymphatics and nerves through the bone.
Interstitial lamellae – irregularly shaped lamellae located between Haversian systems.
23
WORKSHEET NO.3b Name ______________________
Group No. ____
Specialized Connective Tissues Date submitted _______________

Professor ___________________
Rating _____________________
A. Types of cartilage tissues

After examining the prepared slides, draw and label the components of cartilage tissues based on the given descriptions in
the procedure.
HYALINE CARTILAGE ELASTIC CARTILAGE FIBROCARTILAGE
Legend: P = perichondrium; YCh = chondrogenic cells; MCh = mature chondrocytes; Mtx = matrix; Lac = Lacuna;
Bld = blood vessels; Iso = isogenous group of chondrocytes; Coll = collagenous fiber; Elas = elastic fibers
Fib = fibroblasts; Row = rows of chondrocytes
24
B. Types of Bone tissues
After examining the prepared slides, draw the components of bony tissues based on the given descriptions in the procedure.
SPONGY BONE COMPACT BONE
Legend: Per = periosteum; End = endosteum; Ost = osteocytes; Lac = lacuna of osteocyte; Trab = trabecula
Mye = myeloid tissue; Lam = lamella; Can = canaliculi; Hav = haversian canal; Otn = osteon;
25
After examining the tissues, describe the type of CT and their main components by completing the table below.
Other sites with the same

Prepared slide CT Type Function
specialized CT type
Provide structural support to the

Tracheal cartilage
trachea
Epiglottis Provide elasticity to the epiglottis.
Allows movement between

Intervertebral disk vertebrae and provide structural
support of great strength.
The interconnecting network of

trabecula provide maximum
Bone: Spongy layer strength for the minimum mass
within medullary cavities of
bones.
Bone: Shaft of long bone
26
4. Muscular and Nervous Tissues
Muscular tissues are groups of contractile cells that generate motile forces through contraction. Each cell has contractile
filaments called myofilaments containing actin and myosin proteins. Muscles are excited as a response to an appropriate
stimulation and they result into contraction. At this point, mechanical energy is produced where the myofilament components
overlap, causing the whole cell to shorten. Muscle contraction provides the movements taking place in the human body such as
those generated voluntarily by the skeletal muscles. It means that the contractile activity is subject to conscious or voluntary
nervous system control. The spontaneous contraction with rhythmic beating by the heart is due to the involuntary muscular
contraction by cardiac muscles. The passage of ingested foods and liquids through the intestine during digestion, and the
flowing blood and lymph through vessels are due to involuntary muscular contractions by the smooth muscles. Involuntary
contraction means that contraction is generated unconsciously and is regulated by the autonomic nervous system.
Due to differences in structural and functional characteristics, muscle tissues are categorized into three types: skeletal,
smooth and cardiac muscles. However, muscle tissues in general are composite tissues, in that, there is a considerable amount
of connective tissues invested in them. A rich supply of blood vessels, are situated in the connective tissues lying between
muscle cells and between bundles of these cells that will provide those essential requirements. In addition, since muscles are
excitable tissues, nerves and other neural elements are situated also in the connective tissues for contraction to occur.
The nervous tissues are components of the brain and spinal cord, the organs of the central nervous system (CNS) and of
a variety of nerves and ganglia which are parts of the peripheral nervous system (PNS). This special category of tissue is
considered highly cellular in that it is made up of nerve cells called neurons, the structural and functional cells of nervous tissues
along with several types of supporting cells or neuroglia. A neuron is designed to have numerous interconnections with other
neurons through the cell’s cytoplasmic extensions, the dendrites and axons which form a very complex system of
communication. Without these cells the body is unable to receive stimuli and conduct impulses to and from all parts of the
body. Neuroglial cells are the much smaller cells that numbers ten-fold more than neurons in the CNS. These cells have
supportive functions for neurons and they do not conduct impulses like neurons.
The axons of most but not all neurons are covered by myelin sheath and form a nerve fiber. Altogether, when nerve
fibers unite into bundles, they form nerves. The peripheral nervous system (PNS) is consist of cranial and spinal nerves which are
organized by connective tissue wrappings. Nerves are responsible for impulse conduction as the myelinated parts of the axon
fibers conduct impulses away from the neuron cell bodies and sends it to effector or target organs or adjacent nervous tissue.
27
1. Recognize the different muscular tissue types and nervous tissues in a set of prepared slides of
different organs of the body.
Materials:
Prepared slides: smooth muscle, esophagus, skeletal muscle, tongue, spinal cord (human), cerebrum, cerebellum
Procedure:
MUSCULAR TISSUES
1. Smooth Muscle
Myofilament components within smooth muscles do not show cross-striations in histological preparations, thus, giving the
cells a smooth appearance. Each cell possesses a single nucleus located centrally at its broadest part.
Take the prepared slide of the esophagus, transverse section. Using the scanner objective, the more thickly muscular
area called muscularis externa is located inferior to the submucosa (Note: near the inner wall or mucosa there is a thin
muscularis mucosa beneath the lamina propria). There are two layers of muscles, each oriented at a different plane in the
muscularis externa.
Inner Circular muscle - the inner layer of muscles having myofibers running around the organ, and are seen longitudinally cut
when the organ is prepared as transverse section. Take note of the shape of each muscle fiber and the location of the
nucleus within each muscle fiber.
Outer Longitudinal muscle – the outer layer of muscles having myofibers running down the entire length of the esophagus
and are seen transversely cut when the organ is prepared as transverse section. Take note of the shape of each muscle fiber
and the location of the nucleus within each muscle fiber.
Draw and label the basic structure of a smooth muscle as they appear longitudinally and transversely.
28
2. Skeletal muscle
Within each myofiber is a collection of myofibrils which are cylindrical bundles of thick and thin myofilaments that give this
type of muscle a striated appearance.
Examine a section of a skeletal muscle under HPO. Notice the arrangement of muscle fibers into bundles. Draw and
label the basic structure of a skeletal muscle as they appear longitudinally and transversely. This can be seen best in the
tongue section as you locate the muscular region of this organ.
3. Cardiac Muscle
This muscle is composed of larger, yet also long and cylindrical individual cells that contain one to two nuclei. Each cell is
joined end to end with another cell through a complex step-like junction called intercalated disk.
Examine a section of the myocardium or heart muscle under HPO. Take note of the arrangement of fibers that run
longitudinally and the location of the nucleus within the muscle fibers. Look for the intercalated disk, which are the
transverse bands within the myocardium which mark the intercellular boundaries. Draw and label the basic structure of a
cardiac muscle as they appear longitudinally and transversely.
C. NERVOUS TISSUES
1. Neurons and neuroglia
Neuroglia are the much smaller cells found in the in the vicinity of the larger neurons that are light-staining.
Look for the multipolar neurons and neuroglial cells found in the following nervous tissues.
A. Neurons and neuroglia in the gray matter area of spinal cord
Using the scanner objective, locate the anterior horn of the gray matter area seen as the symmetrical H-shaped or
butterfly structure found at the inner layer of the spinal cord c.s. The anterior horn is the more prominent structure
located at the ventral side of the cord. Focus on the large and somewhat stellate-shaped neurons and use a higher
magnification (LPO-HPO) to view this cell.
B. Neurons and neuroglia in the gray matter area of cerebral cortex – these are pyramid-shaped cells.
Using the scanner objective, locate the gray matter area in the cerebrum. The gray matter region lies exterior with
white matter underlying it. The cortex is the outer layer in every gyrus (gyri, plural), the folds in the cerebrum and the
gray matter is seen as the more intensely stained region that lies exterior compared to the less intensely stained internal
29
white matter region. Look for the larger pyramid-shaped neurons and use a higher magnification (LPO-HPO) to view the
cells.
C. Neurons and neuroglia in the gray matter of cerebellar cortex - there are large, flask shaped cells with ramified dendrites
called Purkinje cells arranged as a single layer of neurons in this tissue.
These multipolar neurons have cell bodies that extend outward but the projections are usually not seen completely in
histologic preparations.
Identify the following structures of a multipolar neuron in the gray matter areas of spinal cord and brain.
Cell body or soma – the area which contains the nucleus, nucleolus, numerous different organelles, and the
surrounding cytoplasm or perikaryon.
Nissl bodies in the cytoplasm – the coarse clumps of basophilic substance in the cytoplasm or perikaryon.
Dendritic processes – cytoplasmic extensions in which the processes are usually not visible in preparations. The are
contains Nissl bodies.
Axon hillock – a funnel-shaped cytoplasmic extension that determines the root of an axon. It is devoid of Nissl
bodies.
Axon – a single, long process that extends from the cell body.
2. Nerve: transverse section

Axons Focus on few bundles of nerve axons or nerve fascicle using HPO and look for the following structures:
1. Connective tissue wrappings:
Epineurium around the whole nerve trunk with blood and lymphatic vessels (vasa nervorum), collagen and
fibroblasts, and fat cells.
Perineurium around each fasciculus of nerve fiber: the site of the blood-nerve barrier. Perineurial cells are tightly
attached.
Endoneurium around each individual myelinated nerve fiber but separated from its Schwann cells by a basal lamina.
30
WORKSHEET NO.4
Name ________________________________
Course/Year & Section ________________
Group No. ____
Muscular and Nervous Tissues Professor ___________________
Rating ___________
A. MUSCULAR TISSUES
1. Smooth Muscle: Slide ____________________(400x)
Transverse section Longitudinal section
Arrangement of Number and location of Structures with smooth

Presence of striations Action
myofibers nucleus per myofiber muscle type
Legend: Myo = myofiber; Nuc = nucleus; Cir = circular muscles; Lon = longitudinal muscles;
31
2. Skeletal Muscle: Slide _____________________(400x)
Number and location of Structures with skeletal

Arrangement of fibers Striations Action
nucleus muscle type
Legend: Myo = myofiber; Nuc = nucleus; Str = striations; End = endomysium
32
3. Cardiac Muscle: Slide ______________________ (400x)
Number and location of Structure with cardiac

Arrangement of fibers Striations Action
nucleus muscle type
Legend: Myo = myofiber; Nuc = nucleus; Str = striations ; Int = intercalated disk
33
B. Nervous Tissues (1000 x)
Gray matter area, spinal cord Gray matter area, cerebral cortex Gray matter, cerebellar cortex
Legend: Soma = neuron cell body; Nssl = Nissl bodies; Nuc = nucleus; Axon; Den = dendrite; AxHLK = axon hillock;
Nglia = neuroglia
34
C. Nerve, transverse section (400x)
Legend: NFas = nerve fascicles; Epi = epineurium; Per = perineurium; End = endoneurium; Mye = myelinated axons;
Axn = axon; Schw = Schwann cells
CONCLUSION:
35
5. Blood and Lymphatic Tissues
Blood as a tissue is derived from the embryonic various parts of the body. Hormones are likewise transported
mesenchyme just like the other connective tissues discussed to designated tissues for the regulation of cellular functions,
in the previous activity. This tissue seems fluid but it is while waste metabolic substances from various cells are
actually due to the low ratio of formed elements or blood conveyed to the excretory organs for removal.
cells to the liquid intercellular substance or blood plasma. The formed elements of the blood in circulation were
Blood tends to coagulate when it is left standing for few basically characterized in stained blood smears on glass
minutes after collection. A yellow liquid called serum is slides which were viewed under the microscope. A well-
separated from the clot which contains the formed elements. prepared blood smear reveals separated cells lying flat on
When upon collection into a heparinized hematocrit tube, the glass slide, which when properly stained, allows
blood is kept uncoagulated and its various components can observation of the cells’ characteristic nuclei and cytoplasmic
be separated through centrifugation. The lower layer (~40- organization. Blood obtained from healthy individuals can be
50% of the blood volume) is red due to the packed used to study the normal appearance of formed elements as
erythrocytes or red blood cells, referred to as hematocrit in well as the normal cell counts per unit volume of blood.
this set up. A second layer which is less dense than the All blood cells are formed in the bone marrows
erythrocytes appear as a whitish or grayish thin layer (1% of through a process called haemopoiesis or hematopoiesis.
the blood volume), referred to as buffy coat. It is found The developing blood cells from the marrow can be smeared
immediately above the hematocrit and is consist of the on glass slides, stained and studied in the same manner
leukocytes or white blood cells. A third layer which covers described above. The pluripotential stem cell in the bone
the buffy coat is so thin that it is unseen by the naked eye. marrow is the primitive stem cell type from which all kinds of
This is the layer of the platelets which are fewer and lighter bloods cells are produced. Influenced by favorable
than the components of the buffy coat. Therefore, the microenvironmental conditions and certain growth factors,
formed elements were sedimented and what appears as a the pluripotential stem cell initially forms daughter stem
translucent, yellowish and viscous supernatant is the plasma. cells. The resulting cells are the myeloid and lymphoid
Blood serves as a transporting vehicle as it runs multipotential stem cells from which progenitor cells are
through blood vessels due to the heart’s pumping action. formed. At this point, the progenitor cells begin to generate
The plasma is the fluid which carries the nutrients absorbed cell lineage of a particular blood cell. The myeloid progenitor
in the intestines or synthesized elsewhere for distribution to cells generate precursor cells (~blasts) such as
36
proerythroblasts (for RBCs), myeloblasts (for granular their relative number is an indication of an infection or
leukocytes), monoblasts (for monocytes) and something abnormal is taking place in the body.
promegakaryoblasts (for thrombocytes or platelets). The Aside from the bone marrow being a site of formation
lymphoid progenitor cell will form lymphoblasts (for B and T of blood cells significant for body’s defense against foreign
lymphocytes). Further processes of differentiation take place substances, there are other sites by which this function take
in each cell lineage until the cell reaches its full development. place. The lymphoid system is composed of the lymphoid
The mature blood cells are then released into the circulation organs and lymphoid cells and is designed for defense
to perform their respective roles in the body. function by resulting into what is called an immune response.
The erythrocytes or the red blood cells are the most Primary lymphoid tissues produce lymphoid cells particularly
abundant of all cell types which possess no organelles but the T and B lymphocytes. The bone marrow and the thymus
are filled with hemoglobin which is primarily for O2 and CO2 are considered primary lymphoid organs because these are
binding during gas transport to and from the tissues of the the sites for the B and T lymphocyte formation/ maturation,
body. The leukocytes or white blood cells are classified into respectively. These organs store, release and confer
granular and agranular types. Granulocytes possess specific competence on the lymphocytes that populate the
granules in their cytoplasm which include the neutrophil, secondary lymphoid organs. The secondary lymphoid tissues
eosinophil and basophil. Agranulocytes are WBCs that lack provide the other cells (plasma cells, macrophages, natural
specific granules in their cytoplasm which include the killer cells and dendritic cells) that either participate directly
monocytes and the lymphocytes. Each leukocyte performs a in defense or help activate the lymphocytes in the immune
particular function in terms of defensive responses against response. These are the lymph nodes, spleen, tonsils,
foreign substances such as infectious agents in the body. A mucosa-associated lymphoid tissues (MALT) and Peyer’s
typical range of cell frequency for these leukocytes is used as patches.
reference for a normal or healthy individual. Any change in
37
Activity 5 Blood and Lymphatic Tissues
1. Identify the different cellular components of the blood by recognizing them in blood smear
preparations under the microscope.
2. Relate the role of blood cells in maintaining a healthy body with their normal values or their
abundance in the tissue.
3. Recognize the histologic features of lymphoid tissues and the roles of their component structures in
immune defense.
Materials:
Prepared slides: human blood smear; spleen, thymus, lymph node and tonsil sections.
Procedure:
A. Human blood smear (peripheral blood)
1. Examine the formed elements of the blood using the OIO of your microscope.
2. Scan the whole smear and recognize the different formed elements:
Red blood cell (RBC)/ erythrocyte – small and round, anucleate cell which appear light salmon pink in Giemsa
staining.
Neutrophil – round cell with lobulated basophilic nucleus (3-4 lobes) and unstained granules.
Eosinophil – round cell with bi-lobed nucleus and few large dark pink granules
Basophil – round cell with s-shaped nucleus and large dark blue to black granules
Lymphocytes – round cells with round nucleus that is densely basophilic while the cytoplasm is just a thin band of
dusky blue color on one side of the nucleus.
Monocytes – larger than any other formed element with nucleus that is somewhat, bean-shaped/kidney-
shaped/horseshoe shaped with deep indentation.
Platelets – anucleated, disk-shaped cell fragments.
38
3. Draw each cellular component. Give a brief description of each blood cell as to its staining reaction, size, relative
number in circulation, nuclear features and their function.
B. Bone Marrow smear

1. Examine the bone marrow smear under low magnification and locate areas where the cells are evenly distributed
and do not overlap. Identify the cell from the smear by using the illustration on the blood cell development
presented in page 76 and the description below. The cells are basically developing blood cells of all types called
myeloid cells:
ERYTHROCYTE LINEAGE (in order)

Proerythroblast (Rubriblast) - precursor cell of the erythroid series with a large rounded nucleus, basophilic
cytoplasm.
Basophilic erythroblast (Prorubricyte) - It is smaller than rubriblast, with a nucleus having coarser chromatin,
densely basophilic cytoplasm.
Polychromatophilic erythroblast (Rubricyte) - Smaller than the prorubricytes with more compact nuclear
chromatin, less basophilic cytoplasm, more hemoglobin content than its mother cell. Polychromatophilic
due to its affinity for both acid and basic dyes (because of their content of hemoglobin and RNA
respectively).
Orthochromatophilic erythroblast (Metarubricyte) - Cytoplasm is distinctly acidophilic owing to increased
hemoglobin content and the nucleus is small and pyknotic. Nucleus is eventually expelled.
Reticulocyte – nuclei of metarubricytes have been extruded, hemoglobin becomes more abundant, with small
number of stained network of polyribosomes which will soon disappear.
Erythrocyte - non-nucleated cell with a circular outline. Cell is filled with hemoglobin. In side view, they appear
dumbbell-shaped because of the biconcave nature of their surfaces.
GRANULOCYTE/ LEUCOCYTIC SERIES

Myeloblast - The progenitor cell of the leucocytic series. The cell has lightly basophilic cytoplasm and large and
rounded nucleus. The chromatin is in the form of moderately coarse interconnected strands. They
constitute 0.3 to 0.5 per cent of marrow cells.
Progranulocyte /promyelocyte - It arises and differentiates from myeloblasts. Large cells with round nuclei having
coarse chromatin. Cytoplasm is basophilic with some azurophilic granules. This cell type constitutes about
4 per cent of marrow cells.
39
Neutrophil development:
Neutrophilic myelocyte - derived from progranulocytes. It is smaller in size, with less basophilic cytoplasm
containing differentiated granules and a nucleus with more compact chromatin.
Neutrophilic metamyelocyte – cells with kidney-shaped nucleus. It differentiates into mature neutrophilic
myelocytes.
Neutrophilic bands – characteristic of immature neutrophils wherein the nuclei are horseshoe- or drumstick-
shaped.
Neutrophil (segmented) – characteristic of a mature cell wherein the nucleus is markedly lobulated. The lobules
may be connected with a thin chromatin but compact thread. Cytoplasm is abundant and there are small
granules that may be inconspicuous.
Eosinophil development:
Eosinophilic myelocyte - derived from myeloblasts. The nucleus is rounded or oval with coarser chromatin than in
the myeloblast. Specific eosinophilic granules already appear in the cytoplasm. This cell is capable of
division.
Eosinophilic metamyelocyte – the cytoplasm still contains eosinophilic granules but the nucleus has turned into
kidney shaped or indented. This cell is no longer capable of cell division.
Eosinophilic band – characteristic of immature eosinophil wherein the nucleus is already horseshoe- or drumstick-
shaped, and there are eosinophilic granules in cytoplasm.
Segmented eosinophil – characteristic of mature eosinophil. The nucleus has become lobulated and the lobes are
connected with thin chromatin threads. There is abundant cytoplasm with eosinophilic granules.
Basophil development:
Basophilic metamyelocyte - derived from basophilic myelocyte. The nucleus is oval to kidney-shaped with
cytoplasm containing basophilic granules. Basophilic myelocytes are scarce and may not be seen in a
single marrow smear preparation. This cell is no longer capable of cell division.
Basophilic band - An immature basophil with a horseshoe-shaped nucleus. There are basophilic granules in the
cytoplasm.
OTHER MYELOID CELLS:

Megakaryocytes - very large cell (50 to 70 μm) with a lobulated (mature) or bean-shaped nucleus (immature).
These cells have a blue cytoplasm with many fine azurophilic granules.
40
Plasma cells (mature) - Ovoid cells with deep blue cytoplasm, a clear perinuclear zone, occasional vacuoles and
an eccentric nucleus with clumped chromatin arranged like a cart-wheel.
Degenerating cell - Often found in bone marrow. They are remnants of damaged corpuscles, megakaryocytes, or
myeloblasts. These are primarily artifacts of a marrow smear preparation.
2. Identify the blood cells illustrated in the worksheet and label the stages that are blank.
C. Thymus section
1. Notice that at low power magnification, the gland is covered by a loose connective tissue called capsule and several
lobes are formed within when the connective tissue called septa extend inwardly to separate each lobule.
Look for the following structures and draw them on the worksheet
Cortical region/ Cortex – the outer region of a lobule with compact cell arrangement that makes it appear as dark
blue staining structure.
Medullary region/ Medulla – the central core of a lobule with less densely arranged cells and eosinophilic
structures that make it appear as light staining.
Thymic corpuscles/ Hassal’s corpuscles – the concentric layers of degenerated epithelial reticular cells which are
eosinophilic. These are found in the medulla and considered a key feature in identification of the thymus
tissue.
2. Draw and label the parts of the thymus. Describe the tissue as to its cellular organization.
D. Lymph node
1. Focus on the lymph node section using the scanner.
Connective tissue :
Capsule – dense layer of collagen fibers invested on the lymph node surface.
Hilum – the thickest layer of connective tissue capsule located at the deepest part of the lymph node’s
indentation.
Trabeculae - a connective tissue layer that extends inward from the convex surface of the capsule, also some
CT trabeculae extend from the hilum.
Subcapsular sinus - a clear space visible just beneath the capsule and separating it from the densely basophilic
cortical region.
41
Lymphatic tissue:
Cortex – Basophillic region due to lymphocytes located underneath the capsule. Formed of a series of closely
packed regularly arranged lymph nodules.
Lymph nodule - closely packed circular aggregates of lymphocytes in the cortex of the node (cortical
follicles/ primary nodules). Lymphatic nodules are temporary structures that may develop, disappear,
and redevelop. The size and number of nodules vary widely.
Germinal center - the lighter-staining area in the center of the lymph nodule (secondary nodule or reaction
center). It varies in size with age, being best developed in childhood. It is a temporary structure like
lymph nodules.
Medulla – an irregular and more eosinophilic inner region in the lymph node.
E. Spleen section
1. Focus on the spleen section using the scanner.
Peritoneum – a simple squamous epithelium that lines the capsule of the spleen.
Capsule – a layer of dense collagenous connective tissue that covers the inner part of the spleen.
Trabeculae – connective tissue fibers that extend from the capsule into the interior of spleen to divide it into
lobes.
White pulp – blue stained regions consist mainly of lymphocytes. It appears as a mass of compactly arranged
lymphocytes surrounding an artery.
Red pulp – less dense reddish area, of which color is imparted by erythrocytes.
Arteries – in cross section, arteries are invariably eccentrically placed in white pulp regions
Arterioles – found in red pulp areas as terminal branches of arterial vessels from the white.
42
WORKSHEET NO.5 Name ______________________
Group No. ____
BLOOD AND LYMPHATIC TISSUES Date submitted _______________

Professor ___________________
Rating _____________________
A. Human Blood - Formed Elements
RBC Neutrophil Eosinophil Basophil Monocyte Lymphocyte Platelets

Components Size Nuclear features Relative number/ml Function
Erythrocytes
Platelets
Granular Leukocytes
1. Neutrophil
2. Eosinophil
3. Basophil
Agranular Leukocytes
1. Monocytes
2. Lymphocytes
43
B. Based on the given descriptions on
developing blood cells, identify the
numbered cells in the illustration.
1. ______________________
2. ______________________
3. ______________________
4. ______________________
5. ______________________
6. ______________________
7. ______________________
8. ______________________
9. ______________________
10. ______________________
Figure 2. Blood cell development

Integrated Publishing – MEDICAL: URL: http://www.tpub.com/corpsman/236.htm
44
C. Thymus
Cortical Region Medullary Region
Thymic/ Hassal’s
Corpuscles
Mag. _________
D. Lymph Node
Capsule Hilum
Subcapsular sinus Trabeculae
Cortex Medulla
Lymph nodule
Germinal center
Mag. _________
45
E. Spleen
Peritoneum Capsule
Trabeculae White Pulp
Red Pulp Arteries
Arterioles
Mag. _________
46
Activity 6 Integumentary System
1. Recognize the difference between thin skin and thick skin areas of the body histologically.
2. Identify the component tissues in the 3 basic regions in the skin namely, epidermis, dermis and
hypodermis.
3. Realize the significance of skin as an organ based on its histologic features.
Materials:
Prepared slides: human brown skin, thin skin and thick skin.
Procedure:
a. Human brown skin: Epidermal layer – Thin skin vs. Thick skin
Focus on the epidermal layer of the skin under the LPO.
Stratum corneum – the outermost layer containing dead and dying cells. The cells are flattened, devoid of nuclei and
other organelles and are filled with keratin.
Stratum lucidum – a thin pale layer of keratin, which the cells have accumulated upon maturity and are seen only
when the stratum corneum is very thick.
Stratum granulosum – layer of cells with basophilic granules.
Stratum spinosum - layer of “prickle cells”, which are large and polyhedral in shape.
Stratum basale – layer of cuboidal or low columnar forms with mitotic figures most frequently observed.
Epidermal ridges –
Compare the thin skin section with the thick skin section.
Recognize the structures in this region and take photomicrograph of this tissue. Put labels on the structures.
47
b. Human brown skin section (Dermis)
Look for the following structures and draw them on the worksheet. Use Low magnification to scan the area.
Dermal papillae – form the characteristic wavy boundary between the epithelium and the connective tissue.
Papillary dermis – fine collagen fibers composing 1/5 part of the entire dermis that is located immediately under the
epidermis. It is heavily invested with blood vessels.
Reticular dermis - the lower 4/5 part of the entire dermis consisting of a dense irregular CT with collagen and
reticular fibers. The following specialized structures are found in this region.
- Sweat glands
- Hair follicle
- Sebaceous gland
Arrector pili muscles – a smooth muscle strip that is attached to the hair follicle.
Blood vessels
Nerves
c. Human brown skin: (Hypodermis)

Look for the following structures and draw them on the worksheet.
Fat/adipose cells – round to oval shaped, large cells that look like empty spaces due to the fats that are removed
upon tissue processing.
Nuclei of fat cells – small, dark staining structures that are eventually pressed against the cell membrane, giving
more area for fats in the cytoplasm.
Fibroblasts – cells with large, ovoid nuclei, prominent nucleoli and basophilic cytoplasm that are found near the
adipose cells.
d. Human brown skin section (sweat gland) - HPO

Scan through the entire section at low magnification and focus on the dermis region. Look for the sectioned tubular
region of the sweat gland. Deeper in the dermis region lies the secretory portion of the sweat gland. The excretory
duct ascends through the upper dermis region and penetrates the cell layer of the epidermis.
Secretory cells – the coiled tubular structure lined with large, columnar cells that are lightly eosinophilic.
48
Myoepithelial cells – thin, spindle-shaped cells that surrounds the secretory cells.
Excretory duct – straight tubular structure lined with deep, staining, stratified cuboidal cells, which are smaller than
those of the secretory cells.
Recognize the structures of a sweat gland and take photomicrograph of it. Put labels on the structures.
e. Human brown skin section (hair follicle)

Look for a hair follicle through the section and magnify this structure.
Connective tissue sheath – outer layer that covers the hair follicle.
External root sheath – next to CT sheath; several layers of cells that is continuous with the epidermis.
Internal root sheath – 2 distinct thin layers of pale and granular cells, respectively which become indistinguishable as the
cells combine with the cells of the hair bulb at the base.
Hair shaft – it is consist of the cuticle cell layer, cortex and medulla.
Cuticle cells – inner layer of cells next to internal root sheath that produce the cuticle.
Cortex and medulla – the keratinized inner cores of the hair shaft.
Hair bulb – the expanded region of the follicle in the dermis of skin.
Matrix – the area in the hair bulb where cells of internal and external root sheaths merge. The cells are found partially
surrounding the upper area of the bulb with an inner core of papillary dermis.
Melanocytes – cells found among mitotically active cells in the matrix that produce melanin pigment.
Papillary dermis – the inner core in the bulb consisting of CT with blood vessels.
f. Human brown skin section (sebaceous gland) - HPO

Look for a hair follicle through the section and you will find numerous sebaceous glands surrounding it. Focus on the
sebaceous glands using the HPO. The sebaceous follicles are aggregates of clear cells that are connected to a duct
that opens into the hair follicle.
Hair follicle – located in the dermis region, oriented at an angle to the surface, where hair cuticle is formed.
Sebaceous gland:
basal cells – the lining of the sebaceous gland. A stratified epithelium that has continuity with the external root
sheath of the hair follicle.
49
secretory cells – the gland itself is acinar in shape, which is characterized by polyhedral cells that enlarge, accumulate
secretory materials and become round.
degenerating secretory cells – cells in the anterior portion of the acinar gland which undergo degeneration, a
process in which the cells become the oily secretory product, called sebum.
duct of the sebaceous gland - a short duct lined with stratified epithelia, where the sebum passes through.
Group Lab. Report:

1. Present the photomicrographed images of the tissues into a printed output on white bond paper.
2. Each figure must have a 5 x 5 “ size and must be properly labeled.
3. In each image, give a summary of the significance of the structures to the functions of the skin.
50
Activity 7a Digestive System
1. Identify the structures in each tissue of the oral cavity and digestive tract.
2. Prepare photomicrographed images of the tissues with labels on properly identified structures.
3. Associate the relative function of each structure in the tissue with the main function of a given organ.
4. Exhibit active participation in a group in preparing a single output for this activity.
Materials:
Prepared slides: human lower lip section, tooth section, tongue section , upper esophagus, stomach sections: fundic region
Procedure:
a. Lip section (lower lip)
Examine the section of the lower lip. Use high power magnification to recognize the structures in every portion of the lip.
Shift to low magnification to get a fuller view of the lip section.
External area of the lip - the outer surface that is made up of skin which passes through a transition with the oral
mucosa.
Vermilion border – the more exposed lip surface characterized as a transition zone between the external (skin) area and
inner oral mucosa. The stratified epidermis is lightly keratinized and with abundant blood vessels and nerves but
devoid of hair follicles, sweat and sebaceous glands.
Oral mucosa – the inner lip surface composed of a thick stratified squamous epithelium.
Oral submucosa – underlying tissue that contains salivary glands and a deeper bulk of circumoral skeletal muscles.
Recognize the structures in the lip section and take photomicrograph image of the tissue. Put labels on the structures.
51
b. Tooth: Undecalcified section
Examine under scanner objective the entire undecalcified section of a tooth.
Crown region – the upper portion that is projecting into the oral cavity.
Tooth root – the lower portion that is embedded in the bony ridge of the jaw.
Enamel – a highly mineralized, translucent layer that protects the crown.
Dentine –outer and inner layer next to enamel that forms the bulk of the crown and root composed of calcified
organic matrix similar to that of bone.
Pulp cavity / dental pulp – central area within the dentine that is made up of specialized supporting tissue containing
many sensory nerve fibers.
Cementum – an amorphous calcified tissue surrounding the outer surface of dentine which is anchored to the fibers of
periodontal membrane.
Recognize the structures in the tooth section. Two photomicrograph image sof the tissue can be taken. Put labels on the
structures.
c. Tongue
Examine each portion of the section to locate the various structures of the tongue. Shift between low power and high
power magnification to identify effectively the structures.
Tongue epithelium – varying thickness of stratified squamous cells lining all surfaces.
Papillae - elevations of the oral epithelium and lamina propria that assume various forms and functions.
Filiform papillae – keratinized epithelium with elevations that are conical in shape. These elevations are quite
abundant and are present over the entire surface of the tongue and do not contain taste buds.
Fungiform papillae – elevations that look like mushrooms in that they have a narrow stalk and a smooth-surfaced,
dilated upper part. Taste buds are contained in elevations but scattered on the upper surface, irregularly
interspersed among the filiform papillae.
Circumvallate papillae – the largest papillae with flattened surface. These elevations are least common and are
distributed in the V region in the posterior portion of the tongue.
Taste Buds – are lightly stained oval bodies beneath epithelial linings of fungiform and circumvallate papillae.
Lamina propria – found beneath the epithelium and is consist of dense, collagenous CT. Lamina propria penetrates the
area of serous and mucous glands and skeletal muscles and is invested with nerves, blood vessels and lymphocytes.
52
Skeletal muscles – interlacing bundles of striated muscles that is massively distributed in the entire inner body o f the
tongue.
Recognize the structures in the tongue section. Two photomicrograph images of the tissue can be taken. Put labels on
the structures.
d. Esophagus: Upper region

Examine the lining or mucosa of the upper region of the esophagus and the structures underlying it. Use the scanner or low
power objective to see the entire region.
Stratified squamous epithelium - characterized by numerous layers of cuboidal to polygonal shaped cells from the base
to the midregion, and flattened non-keratinizing cells on the surface.
Lamina propria – structure underlying the epithelia, consist of loose connective tissue rich in blood and lymph vessels
and smooth muscle cells, sometimes also containing glands and lymphoid tissue.
Muscularis mucosa – a thin layer of smooth muscle tissue beneath mucosa consisting of an inner circular and outer
longitudinal muscle fibers.
Submucosa – a region that is composed of collagenous and adipose CT that binds the mucosa to the main bulk of the
muscular wall.
Esophageal glands – small groups of mucus-secreting glands in the submucosal region.
Muscularis externa / propria – a layer of striated muscles that are found only in the upper and middle regions of the
esophagus.
Recognize the structures of the esophageal mucosa, submucosa and muscular regions and take a photomicrograph of the
tissue. Put labels on the structures.
e. Stomach section: Fundus

Focus on the mucosa and submucosa of the fundic region using the HPO. You will find the surface epithelium consisting of
columnar cells that extends into the gastric pits. Draining into the gastric pits are the gastric glands. In the deeper
region, the gastric glands may branch which may appear as transverse and oblique sections.
Mucous neck cells- have round, basal nuclei.
Parietal cells - stain uniformly acidophilic (pink), sometimes pale staining. These are acid secreting cells.
Chief cells/ zymogenic cells – darker staining cells which are basophilic.
53
Three regions of the gastric gland:
(1) Isthmus, lined by surface epithelial cells and parietal cells.
(2) Neck, composed mostly of mucous neck cells and some parietal cells
(3) Base or fundus, composed predominantly of chief cells (zymogenic) and a few parietal cells.
Muscularis mucosa - a thin layer of smooth muscle tissue in the beneath the mucosa consisting of an inner circular and
outer longitudinal muscle fibers.
Submucosa – a region that is composed of collagenous and adipose CT that binds the mucosa to the main bulk of the
muscular wall.
Muscularis externa / propria – an inner region of smooth muscle bundles with an inner circular layer that is reinforced by
an inner oblique layer and an outer longitudinal layer.
Recognize the structures of the fundic mucosa, submucosa and muscularis externa. Take photomicrograph images of these
regions. Put labels on the structures.
Group Lab. Report:

3. In each image, give a summary of the significance of the tissue structures to the functions of the organs in the digestive
system.
54
Activity 7b Digestive System
1. Identify the structures in each tissue of the digestive tract and accessory organs.
2. Prepare photomicrographed images of the tissues with labels on properly identified structures.
3. Associate the relative function of each structure in the tissue with the main function of a given organ.
4. Exhibit active participation in a group in preparing a single output for this activity.
Materials:
Prepared slides: salivary gland, small intestine, large intestine, appendix, rectum, Liver, pancreas
Procedure:
a. Small intestine (Duodenum), c.s.
Observe the uppermost structure that faces the lumen. You will find the fingerlike projections or villi. Notice that some of
these villi would be transeversely cut that you will see rounded structures. The core of each villus is the lamina propria.
Focus on the lower area of the villi where the intestinal glands (crypts of Lieburkϋhn) are found using the HPO. It extends
through the more of lamina propria and to the muscularis mucosae. The gland opens through the intervillous space.
MUCOSA
Villus - fingerlike or leaflike projections, covered by simple columnar epithelium with striated/brush border.
Goblet cells – pale staining cells interspersed among the columnar cells.
Lamina propria – connective tissue underlying the epithelium, which forms the core of each villus and extends beneath
the intestinal glands.
Lacteal – empty lymphatic vessel with very thin endothelium in the lamina propia.
Intestinal gland – or Crypts of Lieberkühn are simple, tubular glands which extend to the muscularis mucosae. The cells
composing this gland are mitotic stem cells, intestinal gland cells, paneth cells, goblet and enteroendocrine cells.
55
Intervillus space – space between individual villi.
Enteroendocrine cells – found interspersed among the cells of the intestinal gland, containing fine granules in the basal
cytoplasm.
Paneth cells – found at the base of the intestinal glands; pyramid-shaped cells with large acidophilic granules and basal
nucleus.
Mitotic cells - found interspersed among the intestinal gland cells usually at the villus tip, with few microvilli. They are
able to divide, migrate, differentiate into other kinds.
Muscularis mucosae – muscular layer underlying the lamina propria consisting of a thin layer of smooth muscles.
SUBMUCOSA, MUSCULARIS, SEROSA

Beneath the structures in the mucosa is a layer of dense irregular connective tissue of submucosa. This region is filled with
glandular structures called Brunner’s or duodenal gland.
Brunner’s / Duodenal gland – pale staining cells arranged as branched tubuloacinar gland. The duct enters the
muscularis mucosae and lamina propria and empties its content to the surface of mucosal intestinal gland (crypt
of lieburkuhn.
Muscularis / muscularis externa – two layers of smooth muscles that surround the duodenal gland.
Inner circular muscle
Outer longitudinal muscle
Serosa – an outer layer of connective tissue that has blood vessels and adipocytes. This structure is covered by simple
squamous epithelium.
Recognize the structures of the small intestine section and take photomicrograph of the mucosa, submucosa and
muscularis. Put labels on the structures. Put labels on the structures.
b. Large intestine section

MUCOSA
Locate the surface epithelia consisting of simple columnar cells, lined with numerous mucus-producing goblet cells and
continues into the intestinal gland (Crypts of Lieberkϋhn). Using the HPO, focus on the deep and straight intestinal
gland area that shows an extension through the lamina propia to the muscularis mucosae.
Columnar cells – surface epithelia, absorptive cells with striated/ brush border.
Intestinal gland – from epithelial ingrowth structure of the large intestine formed as long, simple tubular glands. The
cells composing the secretory portion are goblet cells that colorless due to mucin contents that get washed off
during the tissue processing.
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Lamina propia – connective tissue underlying the epithelia.
Muscularis mucosae - muscular layer underlying the lamina propia consisting of a thin layer of smooth muscles.
SUBMUCOSA, MUSCULARIS, SEROSA

Submucosa – the layer of dense irregular connective tissue beneath muscularis mucosae which has a network of blood
vessels, adipocytes and lymphoid cells.
Muscularis / muscularis externa – two layers of smooth muscles that surround the submucosa.
Inner circular muscle
Outer longitudinal muscle
Serosa – an outer layer of connective tissue that has blood vessels and adipocytes. This structure is covered by simple
squamous epithelium.
Recognize the structures of the large intestine section and take photomicrograph of mucosa, submucosa and muscularis.
Put labels on the structures.
c. Rectum section
Recognize the structures of the rectum section which are similar to the structures of the large intestine and take
photomicrograph of mucosa, submucosa and muscularis. Put labels on the structures.
d. Salivary gland section (sublingual and submandibular)

Scan through the section and focus on one lobule with secretory cells using the HPO. Recognize the serous and mucous
acini and the mixed acini.
Serous acini – smaller and darker-staining pyramidal cells with spherical basal nucleus and apical secretory granules.
Mucous acini – larger than the serous acini, cells are columnar with pale or almost colorless cytoplasm.
Mixed acini – (serous and mucous), mucous acini that are surrounded or capped by one or more serous cells, forming a
crescent-shaped serous demilune.
Myoepithelial cells – thin, spindle-shaped cells that surround the mixed acini.
Duct system where the secretory products pass through:
Intercalated duct – the tube into which the secretory units merge and drain their secretion. It is narrow and still
lined by smaller secretory cells that are cuboidal.
Striated ducts – when intercalated ducts join to form wider ducts; the cells lining this tube has striated
appearance.
57
Interlobular excretory duct – when striated ducts in each lobule converge to form wider ducts. Interlobular ducts
run in the connective tissue
Recognize the structures in the salivary gland section and take photomicrograph of mucosa, submucosa and muscularis.
Put labels on the structures.
e. Pancreas section
Focus on one lobule and recognize the serous acini that surround the pancreatic islets (of Langerhans) using the HPO.
Look for the following structures.
Serous acini – pyramid-shaped cells arranged around small lumina.
Centroacinar cells – light-staining cell found at the lumina or center of the serous acini.
Pancreatic islet – pale-staining cells that are arranged in cord or clumps.
Blood vessels – capillaries that permeates the pancreatic islets
Interlobular connective tissue – CT that separates lobules.
Intercalated duct – smallest duct lined with simple cuboidal epithelium.
Interlobular duct – simple cuboidal cells that become taller and stratified.
f. Liver section
Focus on one hepatic lobule using the scanner objective. This polygonal structure is a unit in the liver section which can be
recognized by a thin connective tissue around it.
Look for the following structures.
Hepatic lobule
Interlobular septa – thin connective tissue that defines one liver lobule.
Portal canals or portal triad – branches of portal vein, bile duct and hepatic artery that are sectioned as a group, found
occasionally at the sides of the hepatic lobule within the interlobular septa.
Central vein – a larger vessel found at the center of hepatic lobules.
Plates of hepatocytes – rows liver cells radiating from the central vein towards the periphery of the lobule.
Sinusoidal lining – blood channels found in between the rows of hepatocytes.
Group Lab. Report:

3. In each image, give a summary of the significance of the tissue structures to the functions of the organs in the digestive
system.
58
8. Central Nervous System
The nervous tissues of the body are components of the run together in a tract. Gray matter has groups of multipolar
brain and spinal cord, which are the central nervous system nerve cell bodies, nerve fibres entering and leaving the gray
and of a variety of nerves distributed as peripheral nervous matter, and preterminal fiber branches (poorly myelinated,
system. This special category of tissue is considered highly hence the gray colour in the fresh, unstained cord). Glial cells
cellular in that it is made up of nerve cells called neurons and blood vessels are in both white and grey matter. Gray
along with several types of supporting cells called neuroglial matter is more vascular. The oligodendrocyte is the principal
cells. The extracellular space represents only 20% of the brain neuroglial cell of white matter. Roots of nerve fibres enter the
and the spinal column. Without these cells the body is unable cord on the dorsal sides; other roots leave on the ventral
to receive stimuli and conduct impulses to and from all parts sides.
of the body. The nervous system as a whole is responsible for The cerebellar and cerebral cortices differ from the
coordinating and controlling many body activities. It spinal cord in these ways:
stimulates muscle contraction, creates an awareness of the (a) gray matter lies to the exterior with white underlying
environment and plays a major role in emotions, memory and it;
reasoning. To do all these things, the cells in nervous tissues (b) tissue of both kinds of cortex is folded into gyri for the
need to be able to communicate with each other by way of cerebral cortex and folia in the cerebellum;
electrical nerve impulses. (c) nerve cells are of various types and are disposed in
The spinal cord is enclosed in connective tissue layers parallel to the pial surface.
meninges with pia extending in at the ventral fissure with the The cerebellar cortex has gray matter consisting of (l)
anterior spinal artery. The ependyma-lined central canal lies Molecular layer (cell processes, but few cells), (2) Purkinje cell
centrally. Surrounding the canal in a butterfly shape is gray layer and (3) Granular cell layer (densely packed small
matter (gray to the naked eye when fresh and unstained). neurons) underlying the white matter, while the cerebral
Horns of gray matter partly separate three columns of white cortex has gray matter consisting of (l) Molecular layer, then
matter: dorsal (posterior), lateral, and ventral (anterior) layers 2, 3, 4, 5, 6 with varying proportions of stellate, fusiform
columns. The white matter is composed of nerve fibers, many and small, medium, and large pyramidal cells underlying white
thickly myelinated, running mainly up or down the cord. matter.
Generally, fibers projecting to or from a particular brain region
59
Activity 8
1. Describe the components of the nervous tissues based on observation using the light microscope.
2. Differentiate the cellular organization of the gray and white matter in the brain and the spinal cord.
3. Recognize the histologic features of different nervous tissues and the roles of their component
structures in the conduction of impulses.
Materials:
Prepared slides: spinal cord, cerebral cortex, cerebellar cortex, spinal ganglion, nerve: transverse and longitudinal section.
Procedure:
A. Cerebral cortex
Use HPO and focus on the cerebral gyri. Look for the following structures:
1. Arrangement of cell bodies of neurons in the gray matter area:
• Molecular layer – relatively few cell bodies chiefly fibers of underlying cells running in many directions but
generally parallel with the surface.
• External granular layer – has a granular appearance; contains cell bodies of many small nerve cells.
• External pyramidal cell layer – pyramid-shaped cell bodies of neurons.
• Internal granular layer – granulate appearance also; contains small cell bodies.
• Internal pyramidal cell layer – pyramidal cells at the internal part; Beth cells – large pyramidal cells in the
motor area of the cerebral cortex.
• Multiform layer – or polymorphic layer; having many cell shapes.
2. White matter (composed of nerve processes) lies below and stains a darker pink. Very little cytoarchitecture is
seen with H&E stain.
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B. Cerebellar cortex
Use LPO and focus on the cerebellar folia. Look for the following structures:
1. Gray matter – has 3 layer of cells:
• Molecular layer – the outer layer contains relatively small neurons.
• Purkinje cells – layer of large, flask shaped cells with ramified dendrites that extend into the molecular layer.
• Granular layer – with abundant small neurons that have intensely stained nuclei.
2. White matter
• The core of each cerebellar folium consisting of myelinated nerve fibers or axons
C. Spinal cord
1. Look for the 3 layers of connective tissue wrappings called meninges:
• Pia mater – the innermost layer with numerous blood vessels
• Arachnoid - the middle layer which constitute a continuous roof over the pia mater
• Dura mater – the outermost layer that is composed of dense connective tissue
2. Gray matter – the structure resembling “H” when seen in cross section; having 2 posterior and 2 anterior horns
(horns are continuous columns). Cell bodies of neurons are located in this region. In the fresh state the tissue is
gray because it contains many cells but not much myelin.
3. White matter – the structure surrounding the “H”-shaped region of the gray matter. Contains many neuroglial
cells which supports the vast numbers of nerve fibers extending up and down the cord. There are no neuronal
cell bodies in this region. In the fresh state the tissue is white because the majority of the fibers are ensheathed
with myelin which is a white fatty material.
61
Name ______________________
WORKSHEET NO.8 Course/Year & Section _________

Group No. ____
NERVOUS SYSTEM Professor ___________________
Rating _____________________
A. Cerebral cortex B. Cerebellar Cortex
Gray Matter:
Molecular cells Gray Matter
External granular cells Molecular cells
External pyramidal cells Purkinje cells
Internal granular cells Granular cells
Internal pyramidal cell White matter
Multiform cells
White matter
Mag. _________ Mag. _________
1. Give the respective functions of the cerebral and cerebellar cortices of the brain.
62
C. Spinal Cord section
Meninges:
Pia mater
Arachnoid
Dura mater
White Matter
Gray Matter
Mag. _________
2. Histologically, how do the gray and white matters of the brain differ from the gray and white matter of the spinal cord?
CONCLUSION
63
Appendix
Histotechnique
Several developments in microscopy required methods in making cellular details more visible and at the same time
preserving tissue structures for long term observation. This method which is basically intended for microscopy is called
“histotechnique”. It initially involves fixation which is the preservation of specimen in a fixative to prevent destruction of tissue
structures due to decay by enzymatic degradation or autolysis and bacterial decomposition. Next will be the removal of water
from the tissue or dehydration. It is followed by clearing process which is the removal of the dehydrant, usually alcohol. These
preliminary procedures are essential and will determine if the tissues can be infiltrated with an embedding medium, which is
paraffin. Melted paraffin enters the tissue crevices and when cooled, tissue blocks can be installed in a cutting device called
microtome. This instrument can make ultra thin sections of the tissue which can be affixed onto the slide. Paraffin is removed
by a solvent, which will then subsequently allow staining of the tissue, usually with two or more dyes. The histological sections
are typically thinner than a single cell. The colors of a prepared tissue are not natural colors, but they make the tissue's
structural details more visible. A widely used stain combination called hematoxylin and eosin, for example, typically colors cell
nuclei purple and the cytoplasm pink.
Materials:
Tissue preparation, fixation and preservation
Live specimen (mice or frog), dissecting set and pan, specimen jar, screw-cap bottles, graduated cylinder, beaker,
distilled water, Chloroform ether, formaldehyde
Dehydration, clearing and paraffin impregnation
Fixed specimen, distilled water, 95% ethyl alcohol, xylene, paraffin, incubator (set at 60 ˚C).
Embedding, Sectioning and affixing sections on slides
Incubator (set at 60 ˚C), paraffin, paper boat casts, metal spatula, alcohol lamp, tissue block cassette, microtome,
64
camel’s hair brush, dissecting needle (blunt-ended), clean glass slides, Haupt’s adhesive, water bath, slide racks,
wooden slide box.
Staining and coverslipping

Coplin jars, utility forceps, xylene, isopropyl alcohol, distilled water, ammonia, hydrochloric acid, filter paper, Harris’
haematoxylin and eosin stains, coverslips (18 x 18 mm), mounting medium (Permount), pipette, drying oven.
Procedure:
A. Smear Preparation of Inner cheek lining cells.
1. Take a sterile toothpick and a clean glass slide.
2. Put a very small drop of water at the center of the slide.
3. Using the blunt end of the toothpick, gently make a slight scraping on the inner lining of the cheek. Be sure you don’t
scrape too much for it might cause bleeding.
4. Mix the scrapings thoroughly with the drop of water on the slide by making a circular motion.
5. Allow the scrapings to spread onto the slide surface at a size similar to a 25 centavo coin.
6. Allow the smear to air dry for several minutes.

7. Stain the smear by following the staining schedule below:
65
Harris’ hematoxylin 3 minutes
Distilled water (using wash bottle) Just enough to rinse
1% Acid alcohol ***
Hematoxylin
staining Tap water 5 dips
Ammonia water (rinse until blue) 1 minute
Distilled water 5-10 dips
Eosin (counterstain) 1 minute
Eosin staining, 95% ethyl 10 dips
dehydration and
clearing Isopropyl – 3 changes 1 minute each
Xylene - 3 changes 1-3 minutes each
8. Mount a coverslip onto the smear using Canada balsam or Permount which are xylene-based. Place a drop of the
mountant on the slide, taking care to leave no bubbles. Angle the coverslip and let fall gently onto the slide. Allow the
mountant to spread beneath the coverslip, covering all the tissue and allow it to dry.
B. Demonstration of histotechnique procedure (Preparation of tissue sections)

1. Tissue preparation, fixation and preservation
Fix cut pieces of organs with 10% formaldehyde in properly labeled jars within 24-48 hours at room temperature. After
12 hours, these have turned firm enough to be trimmed into appropriate size which is at least 5-7 mm thick and with an area
of about 10 mm x 20 mm. Further fix the tissue pieces until 48 hours.
2. Dehydration, clearing and paraffin impregnation

Dehydration is the removal of water from the tissue. Replace water with a series of increasing grades of alcohol from
70% to 95% to 100%, following the schedule given below.
Clearing is the removal of the dehydrant (alcohol). Replace the absolute alcohol with xylene, the most common
clearing reagent that will be miscible with the embedding medium (paraffin).
66
Impregnation or infiltration is the gradual entry of the embedding medium into the tissue. Impregnate the tissue with
melted paraffin by using 2 changes of melted soft paraffin (42-50 ̊melting point), followed by two changes melted hard
paraffin (58-60 ̊melting point). This procedure is carried out at 60˚C inside a hot air oven or in a water bath.
Follow the schedule of a manual tissue processing below.
Manual Tissue Processing:

STAGE REAGENT DURATION (hrs)
70% ethanol 12
95% ethanol 12
Dehydration Isopropyl 1 6
Isopropyl 2 6
Isopropyl 3 6
Xylene 1 4
Clearing Xylene 2 4
Xylene 3 4
Melted paraffin – Soft 3
Paraffin
impregnation Melted paraffin – Soft 12
(inside 60˚C hot air Melted paraffin – Hard 2
oven or incubator)
Melted paraffin – Hard 2
TOTAL TIME 73 hours ( ~ 3 days)
Gently shake the jar with pieces of organs from time to time in every reagent to facilitate the entry of liquids into the tissue.
67
3. Embedding, Sectioning and affixing sections on slides
Place the paraffin-impregnated tissues into casts, which are sometimes prepared as a small paper box or as a thin
plastic square mold. Properly orient or align the tissues in the cast and pour the molten paraffin over them. Allow this warm
paraffin with tissue to cool and solidify into what is called a paraffin-tissue block, which is removed from its cast.
Attach the paraffin-tissue block onto the block cassette using extra paraffin that is melted with a warmed metal
spatula. This is in preparation for microtomy, the cutting of tissues into about 4-8 microns (μm) in thickness using the
microtome. Set the paraffin-tissue blocks onto a cold surface (4˚C) to harden the face or side to be cut.
Trim the attached paraffin tissue block with a sharp blade, making the sides parallel and atleast 2-3 mm of paraffin
should surround the tissue. Install the microtome knife or blade in place and set the correct clearance angle. Fit the trimmed
block into the block holder of the microtome and adjust so the edge offering least resistance meets the knife edge first.
Advance the block until it just touches the knife. Course cut the block at 15 μm by rotating the flywheel until the full face has
been trimmed. Set the advance feed dial to 6 μm, which is the desired thickness for most purposes. Cut a series of sections,
which form what is called a ribbon. Separate the ribbon from the knife edge using a moist camel hair brush. Once the knife
has advanced to its full extent, a blinking of a small, red light reminds the user that it has to be returned back to its starting
position by rotating the reverse feedwheel.
Place the sections or ribbons on a flat board lined with a white or colored paper and use a sharp blade to separate one
section from a ribbon. Float the section on a warm water bath so that wrinkles along with some air bubbles trapped beneath
the paraffin will be removed. The temperature of the water should be approximately 10˚C below the melting point of the
paraffin used in the block.
Prepare a clean glass slide and apply a very thin coat of albumin or Haupt’s adhesive on the surface using your pinky
finger. Use this adhesive-coated slide to pick up the expanded section from the water bath. Hold the slide vertically and
mostly beneath the surface of the water. The section must be apposed to the slide so that when it is lifted from the water, it
draws the section with it. Dry the slide overnight at room temperature or on a hot plate or hot air oven set just above the
melting point of the paraffin.
4. Staining and coverslipping

Reverse the embedding process in order to get the paraffin wax out of the tissue that will allow water-soluble dyes to
penetrate the sections. Before doing the staining process, run the slides through xylenes to alcohols to water, in order to
deparaffinize and rehydrate the tissues. Tissues will not be stained when there is paraffin. The routine stain is that of
hematoxylin and eosin (H and E).
68
Follow the schedule of H. and E. staining procedure below.
Hematoxylin and Eosin staining:
STAGE REAGENT TIMING

Xylene – 3 changes 3 minutes each
100% Isopropyl – 2 changes 1 minute each
Deparaffinize and
rehydrate 95% Ethyl 5-10 dips
sections
70% Ethyl 5-10 dips
Harris’ hematoxylin 5-10 minutes
Distilled water (using wash bottle) Just enough to rinse
Tap water 5-10 dips
Hematoxylin 1% Acid alcohol ***
staining
Tap water 5 dips
Ammonia water (rinse until blue) 1 minute
Eosin (counterstain) 2 to 5 minutes
Eosin staining, 95% ethyl 15 dips
dehydration and
clearing Isopropyl – 3 changes 1 minute each
Xylene - 3 changes 1-3 minutes each
*** Differentiate sections in 1 % acid-alcohol and then wash well in water. Repeat if more stain needs to be removed. This step requires microscopic control
to ensure that only nuclei are stained.
69
The stained slide is taken through a series of alcohol solutions to remove the water, then through clearing agents to a
point at which a permanent resinous substance beneath the glass coverslip, or a plastic film, can be placed over the section.
This is done to accomplish three things: to protect the tissue from being scratched, to provide better optical quality for
viewing under the microscope, and to preserve the tissue section for years to come.
Coverslip slides with Canada balsam or Permount.
D. Clean the area over and around the mounted coverslips with cotton moistened with xylene. Label the prepared slide
accordingly.
HISTOLOGY
FROG
Liver section
H. & E.
HUB 31 Grp.2A 1st
Sem.
SY 2010-2011
E. Evaluation of prepared slides (selected slides with faults)

Carefully study the tissue slides prepared by the group and compare it to the set of slides distributed by the instructor
(preferably from the same organ and species source). Identify the tissue “faults” due to tissue processing.
70
References:
BOOKS:
Cormack, D.H. 1987. HAM’S HISTOLOGY 9th Edition. J.B.Lippincott Company. Philadelphia USA
Gartner, L.P., Hiatt, J.L., Strum, J.M. 2003. Board Review Series CELL BIOLOGY AND HISTOLOGY 4th Edition. Lippincott Williams
and Wilkins. Philadelphia USA
Eroschenko V. 2003. DI FIORE’S ATLAS OF HISTOLOGY WITH FUNCTIONAL CORRELATION. William and Wilkins Baltimore USA
Junqueira, L.C., Carneiro, J. 2005. BASIC HISTOLOGY TEXT AND ATLAS, 11th Edition. McGraw-Hill Publishing. USA
Young, B.B., Heath, J.W. 2000. WHEATER’S FUNCTIONAL HISTOLOGY. Elsevier Limited. Churchill Livingstone
INTERNET SOURCES:
Beresford, W.A. HISTOLOGY FULL TEXT (Web version). http://wberesford.hsc.wvu.edu/histol.htm
Bergman, R.A., Afifi, A.K., Heidger Jr, P.M. (Editors). 2008. ATLAS OF MICROSCOPIC ANATOMY - A Functional Approach:
Companion to Histology and Neuroanatomy: Second Edition.
http://www.anatomyatlases.org/MicroscopicAnatomy/MicroscopicAnatomy.shtml
Caceci, T. 1998. VM8054: Veterinary Histology Version 2. LABORATORY EXERCISES.
http://education.vetmed.vt.edu/curriculum/vm8304/lab_companion/histo-path/VM8054/Labs/labtoc.htm
Caprette, D.R. 1995 (Updated 2005) Light Microscopy EXPERIMENTAL BIOSCIENCES Introductory Laboratory 211 Rice University
http://www.ruf.rice.edu/~bioslabs/methods/microscopy/microscopy.html
Roy, E.C. 2007 LABORATORY HISTOLOGY. http://www.adam.com.au/royellis/
Slomianka, L. 2006. BLUE HISTOLOGY – Notes http://www.lab.anhb.uwa.edu.au/mb140/
Helpful Links:
Histology World. 2007. http://www.histology-world.com/
Histology Lab Manual. 2008. SUNY Downstate Medical Center. http://ect.downstate.edu/courseware/histomanual/
71

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Histology

II. Care of the microscope

C. Human squamous epithelium (inner-cheek lining cells)

D. Pathologic tissue sections

THE CELL: STRUCTURE AND FUNCTION Date submitted _______________

Cell membrane Cytoplasm

Nucleus Cytoplasmic extension

Neuron cell body

Cell membrane Cytoplasm

Nucleus Chromatin materials

Nucleus Chromatin materials

Nucleoplasm Cell membrane

STRUCTURE SPECIAL CHARACTERISTICS FUNCTION

c. nuclear sap/ nucleoplasm

b. Kidney section: Renal tubules of the cortical parenchyma.

c. Gall bladder: Fingerlike folds of the mucosa

1.2. STRATIFIED OR COMPLEX TYPE

b. Parotid gland: Larger excretory duct/ interlobular ducts

c. Parotid gland: Main excretory ducts/ interlobar ducts

d. Trachea: Inner Lining

e. Urinary bladder: Lining of the mucosa

Function Function Function Function

STRATIFIED SQUAMOUS STRATIFIED CUBOIDAL STRATIFIED COLUMNAR TRANSITIONAL

Function Function Function Function

CONNECTIVE TISSUES (CT)

Adipocytes of brown, multilocular adipose tissue contain many lipid droplets.

b. Human brown skin: Hypodermal region

c. Lymph node: Stroma

d. Human brown skin: Dermal region

Loose and Dense Connective Tissues Group No. ____

A. CONNECTIVE TISSUE components

COLLAGEN FIBERS ELASTIC FIBERS RETICULAR FIBERS FIBROBLASTS

LYMPHOCYTES PLASMA CELLS MACROPHAGES ADIPOCYTES/ FAT CELLS

Other sites with the same CT

b. Human brown skin

Reticular CT serves as fine

d. Dermis region of the

e. Tendon section A source of great tensile

a. Trachea: Cartilage plate

c. Intervertebral disc: Fibrocartilage

d. Bone section: Spongy portion

e. Bone, decalcified: Shaft of a long bone

Specialized Connective Tissues Date submitted _______________

A. Types of cartilage tissues

HYALINE CARTILAGE ELASTIC CARTILAGE FIBROCARTILAGE

SPONGY BONE COMPACT BONE

Other sites with the same

Provide structural support to the

Epiglottis Provide elasticity to the epiglottis.

Allows movement between

The interconnecting network of

Bone: Shaft of long bone

2. Nerve: transverse section

1. Smooth Muscle: Slide ____________________(400x)

Transverse section Longitudinal section

Arrangement of Number and location of Structures with smooth

Transverse section Longitudinal section

Number and location of Structures with skeletal

Legend: Myo = myofiber; Nuc = nucleus; Str = striations; End = endomysium

Transverse section Longitudinal section

Number and location of Structure with cardiac

B. Bone Marrow smear

ERYTHROCYTE LINEAGE (in order)

Mag. _ Mag. _