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Examination
Dr.dr I Wayan Putu Sutirta Yasa, M.Si
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Structure of hemoglobin
“Globin" consists of four polypeptide chains:
two alpha chains, each with 141 amino
acids and two beta chains, each with 146
amino acids.
The α and β globin chains are very similar in
structure and each one of them is linked
with a heme molecule. Each heme group
can combine with 1 molecule of oxygen or
CO2.
A heme group is a flat ring molecule
containing carbon, nitrogen and hydrogen
atoms, with a single Fe2+ ion at the center.
Without the iron, the ring is called a
porphyrin.
Theoxygen-binding site of Hb is the heme
pocket present in each of the four
polypeptide chains; a single atom of
oxygen forms a reversible bond with the
ferrous iron at each of these sites, so a
molecule of Hb binds four oxygen
molecules; the product is oxyhemoglobin
(O2Hb).
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HEMOGLOBIN FUNCTION
The oxygen delivery function of Hb, that is its ability
to "pick up" oxygen at the lungs and "release" it to
tissue cells is made possible by minute
conformational changes in quaternary structure that
occur in the hemoglobin molecule and which alter
the affinity of the heme pocket for oxygen. Hb has
two quaternary structural states: the deoxy state
(low oxygen affinity) and the oxy state (high oxygen
affinity).
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A range of environmental factors determine
the quaternary state of Hb and therefore its
relative oxygen affinity. The
microenvironment in the lungs favors the oxy-
quaternary state, and thus Hb has high
affinity for oxygen here.
5
By contrast, the microenvironment of the
tissues induces the conformational change
in Hb structure that reduces its affinity for
oxygen, thus allowing oxygen to be
released to tissue cells.
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What is a Hemoglobin Test?
A hemoglobin test measures the levels of hemoglobin in your blood.
Hemoglobin is a protein in your red blood cells that carries oxygen from your
lungs to the rest of your body. If your hemoglobin levels are abnormal, it may
be a sign that you have a blood disorder.
What is it used for?
A hemoglobin test is often used to check for anemia, a condition in which your
body has fewer red blood cells than normal. If you have anemia, your cells
don't get all the oxygen they need. Hemoglobin tests are also frequently
performed with other tests, such as:
Hematocrit, which measures the percentage of red blood cells in your blood
Complete blood count, which measures the number and type of cells in your
blood
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Why do I need a hemoglobin test?
plan.
Higher than normal results
If your hemoglobin level is higher than normal, it may be the result of:
Polycythemia vera — a blood disorder in which your bone marrow makes too
many red blood cells
Lung disease
Dehydration
Living at a high altitude
Heavy smoking
Burns
Excessive vomiting
Extreme physical exercise
If you've been previously diagnosed with polycythemia vera, an elevated
hemoglobin level may indicate a need to alter your treatment plan.
If your hemoglobin level is below or above normal, your doctor may want to
evaluate the hemoglobin test results along with those of other tests, or 13
additional tests may be necessary, to determine next steps.
If any of your levels are abnormal, it does not
necessarily indicate a medical problem needing
treatment.
Diet, activity level, medications, a women's menstrual
cycle, and other considerations can affect the results.
In addition, you may have higher than normal
hemoglobin if you live in a high altitude area. Talk to
your health care provider to learn what your results
mean.
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Results
15
Why it's done
You may have a hemoglobin test for several reasons:
To check your overall health. may test your hemoglobin
as part of a complete blood count during a routine
medical examination to monitor your general health and
to screen for a variety of disorders, such as anemia.
To diagnose a medical condition. may suggest a
hemoglobin test if you're experiencing weakness, fatigue,
shortness of breath or dizziness. These signs and
symptoms may point to anemia or polycythemia vera. A
hemoglobin test may help diagnose these or other medical
conditions.
To monitor a medical condition. If you've been diagnosed
with anemia or polycythemia vera, may use a hemoglobin 16
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Sahli's Hemoglobinometer
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Materials of the experiment
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Procedure Sahli's test
3- Put the pipette into HCL solution and suck up a
little of the acid and let it flow down in the
mixture.
4- By the pipette Suck up a little destilate water
and add to the mixture. Repeat 2 to 3 times for
washing the pipette.
5- The mixture after about 2 minutes will be dark
brown and clear.
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Procedure Sahli's test
6- Add dist water drop by drop, mixing well by a glass rod and
compare the color of this tube with the standard tube.
7- After 5 minutes, read the haemoglobin content from the scale
on the graduated mixing tube.
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Result
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less room lighting
less room lighting
less room lighting
KCN Cyanmethemoglobin
Cyan-Methemoglobin
0 0 0 5
5 0.125 1 4
10 0.250 2 3
15 0.375 3 2
7.5 0.188 2.5 2.5
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Procedure
30
Standard Curve calculation
∆Y
∆X
Concentration of Standard 31
Calculation
32
Discussion
34
The normal ranges
Newborns 17-22gmdl
One (1) week of age 17-22gm\dl
One (1) week of age 15-20gmdl
One (1) month of age 11-15gm\dl
Children 11-13gm\dl
Adult men 14-18gm\dl
Adult women 12-16gm\dl
Men after middle age 12.4-14.9gm\dl
Women after middle age 11.7-13.8gm\dl
• Normal values in an adult are 12 to 18 grams per deciliter
(100 milliliters) of blood. 35
1. Strengths of the Cyanmeth Method
a. Accurate inspection.
b. Reagent and a device for measuring hemoglobin levels can be controlled
with a stable standard solution.
2. Lack of the Cyanmeth Method
a. Tools for measuring absorbance (spectrophotometers or photometers)
are expensive and require electricity
a. Reagent drafkin solution containing cyanide, is toxic
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37
Acuan
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39
Hb examination according to Sahli was
classified
into the colorimetry
Basic consept method
In principle, blood Hb is converted to Hematin chloride,
whose color becomes dark brown (tengguli).
the color that occurs is diluted with aquadest to the standard col
of Hematin chloride
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Instrument : HAEMOMETER"
1, A pair of glass cylinders contain a standard color solution,
we call it a color comparator.
2, Measuring tube (thinner tube) which has lines,
a scale that shows Hb level, The lowest scale is number 2.
3, Capillary blood pipette (Hemoglobin Pipette)
about the size that has a volume of 20 mm3 on the boundary line.
4, Pipette pasteurs for distilled water
5, Stirring glass stems
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Material : HCl 0,1N and Aquadestilata
Examination Method :
1, Prepare the tube and fill it with 0.1 N HCl to the lowest line (in
number 2)
2, With a pipette Hb suction of blood until the number 20 cm do not let
there be air bubbles who was sucked in
3, Pour blood into the diluent tube, rinse with HCl if there is still blood
in the pipette
4, Leave for 1 minute
5, Add the aquadest drop by drop, stirring with a stirring glass rod
6, Compare the solution in the diluent tube with the standard solution
color
7, the color is the same the aquadest addition is stopped, read the Hb
level on 42
43
Bahan
Reagen yang diperlukan : HCl 0,1 N dan Aquadestilata
Cara Pemeriksaan
1. Siapkan tabung dan isilah dengan HCl 0,1 N hingga garis yang terendah (pada angka 2).
2. Dengan pipet Hb hisap darah sampai angka 20 cmm jangan sampai ada gelembung udara
yang ikut dihisap
3. Tuang darah ke dalam tabung pengencer, bilas dengan HCl bila masih ada darah dalam pipet
4. Biarkan selama 1 menit
5. Tambahkan aquadest tetes demi tetes, aduk dengan batang kaca pengaduk
6. Bandingkan larutan dalam tabung pengencer dengan warna larutan standart
7. Bila sudah sama warnanya penambahan aquadest dihentikan, baca kadar Hb pada skala yang
ada di tabung pengencer.
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