UNDERSTANDING THE

PRINCIPLES OF
INSTRUMENTATION
LEARNING ACTIVITY NO. 19
INTRODUCTION
• Instrumentation has been the bridge of convenience for the
medical technologist inside the clinical laboratory. Since there
is a need for quantitative assessment, instrumentation has
been utilized over the years for a more effective patient care
system.
• The methodologies used in technologically sophisticated
automated analyzers are based on traditional approaches and
techniques.
INTRODUCTION

• There are general methods for most automated and manual assays
in the chemistry laboratory, which includes the use of
spectrophotometry, ion-selective electrodes, electrophoresis,
nephelometry, and immunoassays.
• These instrumentation techniques, and their correct application,
serve as the backbone of good quality results generation thus
making it highly important for the medical laboratory scientist to
be equipped with such a knowledge, skill, and attitude.
WHAT IS ELECTROMAGNETIC RADIATION?

• It refers to the waves of the electromagnetic field, propagating

through space, carrying electromagnetic radiant energy. It is a
form of energy that is all around us and takes many forms, such as
radio waves, microwaves, infrared, light, ultraviolet, X-rays, and
gamma rays.
COLORS OF THE VISIBLE SPECTRUM AND THE
RADIANT ENERGY BY THE INVISIBLE SPECTRUM
BEER’S LAW

• Beer's Law is also known as the Beer-Lambert Law, the Lambert-

Beer Law, and the Beer–Lambert–Bouguer law.
• Beer's law states that the concentration of a chemical solution is
directly proportional to its absorption of light.
• The premise is that a beam of light becomes weaker as it passes
through a chemical solution. The attenuation of light occurs either
as a result of distance through solution or increasing
concentration.
EQUATION FOR BEER’S LAW

• A = εbc

• where A is absorbance (no units)

• ε is the molar absorbtivity with units of L mol-1 cm-1
(formerly called the extinction coefficient)
• b is the path length of the sample, usually expressed in cm
• c is the concentration of the compound in solution,
expressed in mol L-1

RHODAMINE 6G
PHOTOMETRY
• Atomic Emission Spectrophotometry - is based on the principle that when a
molecule is applied energy in the form of light or heat, the molecules get
excited and move from lower energy level to higher energy level. At this state
the molecules are unstable. Therefore, the excited molecule jumps from
higher energy level to lower energy level, emitting radiation. The radiations
are emitted in the form of photons. The wavelengths of photons emitted are
recorded.
• Atomic absorption spectroscopy - is based on the principle that when a beam
of electromagnetic radiation is passed through a substance, the radiation may
either be absorbed or transmitted depending upon the wavelength of the
radiation.
• Molecular Luminescence Spectroscopy - the detection of small amounts of
impurities in naturally occurring solutions such as rivers and groundwater.
NON-ABSORPTIVE METHODS
• Nephelometry – light is passed through the sample solution (suspended
particles) directly and the amount of scattered radiation is measured
generally at 90°C.
• Turbidimetry - It is performed by measuring the turbidity in a water sample by
passing light through the sample being measured.
• Refractometry – It is used to measure of the speed of light through the
substance, usually liquid.
• Osmometry – It is used to measure hydration levels using the osmolality of
serum or urine.
ELECTROCHEMISTRY
A. Potentiometry
• pH electrode – an electrode that measures the concentration of hydrogen
ions.
• pCO2 electrode – An electrode that measures carbon dioxide (CO2).
• Ion-selective electrode - A transducer that converts the activity of a specific
ion dissolved in a solution into an electrical potential. Consists of a thin
membrane across which only the intended ion can be transported. The
transport of ions from a high conc. to a low one through a selective binding
with some sites within the membrane creates a potential difference.
OTHERS
• Electrophoresis - is a technique used to separate and sometimes purify
macromolecules - especially proteins and nucleic acids - that differ in size,
charge or conformation. As such, it is one of the most widely-used techniques
in biochemistry and molecular biology.
• Chromatography – It is based on the principle where molecules in mixture
applied onto the surface or into the solid, and fluid stationary phase (stable
phase) is separating from each other while moving with the aid of a mobile
phase.
• Gas Chromatography/Mass Spectrometry (GC/MS) - separates chemical
mixtures (the GC component) and identifies the components at a molecular
level (the MS component). It is one of the most accurate tools for analyzing
environmental samples. The GC works on the principle that a mixture will
separate into individual substances when heated.