JIMMA UNIVERSITY

INSITITUTE OF HEALTH
FACULTY OF HEALTH SCIENCE
COLLEGE OF PUBLIC AND MEDICAL SCIENCES
SCHOOL OF MEDICAL LABORATORY
DEPARTMENT OF MEDICAL LABORATORY
AND PATHOLOGY
ASSIGMENT OF INSTRUMENTATION

BY: DAMISE BACHA ID NO: RU3225/10

SUBMITTED TO: SINTAYEHU ASAYE (MSC)

DATE OF SUBMITTION…………..NOV 4, 2019

JIMMAA, ETHIOPIA

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Table of contents…………………………………………………..page number

 Acknowledgement…………………………………………………………………..3
 Introduction……………………………………………………...3
 Chromatography………………………………………………………………………4
 What is chromatography...........................................................4
 Classification of chromatography……………………………………………..5
 Application of chromatography…………………………………………………9
 Calibration and standards…………………………………………………………11
 Radiochemistry…………………………………………………………………………12
 Determination of radionuclide…………………………………………………12
 Application of radiochemistry………………………………………………….13
 Radiometry……………………………………………………………………………..13
 What is radiometry…………………………………………………………………13
 Conclusion……………………………………………………………………………..15
 Reference……………………………………………………………………………….15

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Acknowledgement

My first and foremost gratitude is forwarded to my almighty God for his support and protection
in all my life.
Next, my heart full thanks goes to my INSTRUCTOR SINTAYEHU ASAYE (MSC) for give as
this assignment and make as to practice on the main important topic.

Introduction This assignment contains mainly three parts those are chromatography its
definition, classification and application; radiochemistry its definition, history and some parts
finally its application at the end there is radiometry its definition, its instruments, units and who
to reveal it.

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 Chromatography
What is chromatography?
Chromatography 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. The factors effective on this separation process include
molecular characteristics related to adsorption (liquid-solid), partition (liquid-solid), and affinity
or differences among their molecular weights. Because of these differences, some components of
the mixture stay longer in the stationary phase, and they move slowly in the chromatography
system, while others pass rapidly into mobile phase, and leave the system faster.

Based on this approach three components form the basis of the chromatography technique.

 Stationary phase: This phase is always composed of a “solid” phase or “a layer of a liquid
adsorbed on the surface a solid support”.

 Mobile phase: This phase is always composed of “liquid” or a “gaseous component.”

 Separated molecules

The type of interaction between stationary phase, mobile phase, and substances contained in the
mixture is the basic component effective on separation of molecules from each other.
Chromatography methods based on partition are very effective on separation, and identification
of small molecules as amino acids, carbohydrates, and fatty acids.

Various chromatography methods have been developed to that end. Some of them include
column chromatography, thin-layer chromatography (TLC), paper chromatography, gas
chromatography, ion exchange chromatography, gel permeation chromatography, high-pressure
liquid chromatography, and affinity chromatography.

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Classification of chromatography

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  Column chromatography

 Ion-exchange chromatography

 Gel-permeation (molecular sieve) chromatography

 Affinity chromatography

 Paper chromatography

 Thin-layer chromatography

 High-pressure liquid chromatography (HPLC)

Column chromatography

Since proteins have difference characteristic features as size, shape, net charge, stationary phase
used, and binding capacity, each one of these characteristic components can be purified using
chromatographic methods. Among these methods, most frequently column chromatography is
applied. This technique is used for the purification of biomolecules. On a column (stationary
phase) firstly the sample to be separated, then wash buffer (mobile phase) are applied. r.

Ion- exchange chromatography

Ion- exchange chromatography is based on electrostatic interactions between charged protein

groups, and solid support material (matrix). Matrix has an ion load opposite to that of the protein
to be separated, and the affinity of the protein to the column is achieved with ionic ties. Proteins
are separated from the column either by changing pH, concentration of ion salts or ionic strength
of the buffer solution

Gel- permeation (molecular sieve) chromatography

The basic principle of this method is to use dextran containing materials to separate
macromolecules based on their differences in molecular sizes. This procedure is basically used to
determine molecular weights of proteins, and to decrease salt concentrations of protein solutions.
In a gel- permeation column stationary phase consists of inert molecules with small pores

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 polyacrylamide are also
used as column materials.

Affinity chromatography

This chromatography technique is used for the purification of enzymes, hormones, antibodies,
nucleic acids, and specific proteins. A ligand which can make a complex with specific protein
(dextran, polyacrylamide, cellulose etc) binds the filling material of the column. The specific
protein which makes a complex with the ligand is attached to the solid support (matrix), and
retained in the column, while free proteins leave the column.

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FIGURE 4

Affinity chromatography.

Paper chromatography

In paper chromatography support material consists of a layer of cellulose highly saturated with
water. In this method a thick filter paper comprised the support, and water drops settled in its
pores made up the stationary “liquid phase.” Mobile phase consists of an appropriate fluid placed
in a developing tank. Paper chromatography is a “liquid-liquid” chromatography.

Thin-layer chromatography

Thin-layer chromatography is a “solid-liquid adsorption” chromatography. In this method

stationary phase is a solid adsorbent substance coated on glass plates. As adsorbent material all
solid substances used. In column chromatography (alumina, silica gel, and cellulose) can be
utilized. In this method, the mobile phase travels upward through the stationary phase the solvent
travels up the thin plate soaked with the solvent by means of capillary action. During this

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procedure, it also drives the mixture priorly dropped on the lower parts of the plate with a pipette
upwards with different flow rates. Thus the separation of analytes is achieved. This upward
travelling rate depends on the polarity of the material, solid phase, and of the solvent.

Gas chromatography

In this method stationary phase is a column which is placed in the device, and contains a liquid
stationary phase which is adsorbed onto the surface of an inert solid. Gas chromatography is a
“gas-liquid” chromatography. Its carrier phase consists of gases as He or N2. Mobile phase which
is an inert gas is passed through a column under high pressure. The sample to be analyzed is
vaporized, and enters into a gaseous mobile phase phase.

Pseudo affinity chromatography

Some compounds as anthraquinone dyes, and azo-dyes can be used as ligands because of their
affinity especially for dehydrogenases, kinases, transferees, and reductases the mostly known
type of this kind of chromatography is immobilized metal affinity chromatography (IMAC).

High-prssure liquid chromatography (HPLC)

Using this chromatography technique it is possible to perform structural, and functional analysis,
and purification of many molecules within a short time, This technique yields perfect results in
the separation, and identification of amino acids, carbohydrates, lipids, nucleic acids, proteins,
steroids, and other biologically active molecules, In HPLC, mobile phase passes through
columns under 10–400 atmospheric pressure, and with a high (0.1–5 cm//sec) flow rate

Applications of Chromatography

Qualitative analysis; the chromatogram of a run provides one piece of information about each species in a
sample, the retention time or position, but this information is important and can be used to identify
multiple species in a mixture. Of course a chromatogram also provides information of the absence of a
species from a sample. For example, lack of steroids from an Olympic weightlifter’s urine sample is seen
as a great benefit to the career of the weightlifter !

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 Quantitative analysis
Chromatography also provides some important qualitative information about the separated species present
in a mixture. For column chromatography, comparison of the height of a peak or the area under a peak
can be compared with standards to determine the amount of that species presentght line and the
perpendicular distance to the peak is measured. Peak broadening can reduce the accuracy of this
approach.

Application areas of chromatography in medicine

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Chromatography technique is a valuable tool for biochemists, besides it can be applied easily
during studies performed in clinical laboratories For instance, paper chromatography is used to,

steroids.

Calibration and standards

The easiest approach is to prepare a series of standard solutions that approximate the composition of the
unknown. The chromatograms of the standards are used to plot the peak heights or areas against
concentration to obtain a standard curve. This should be a straight line that passes through the origin.

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 .

Radiochemistry

Radiochemistry is the chemistry of radioactive materials, where radioactive isotopes of elements are used

to study the properties and chemical reactions of non-radioactive isotopes (often within radiochemistry
the absence of radioactivity leads to a substance being described as being inactive as the isotopes

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are stable). Much of radiochemistry deals with the use of radioactivity to study ordinary chemical
reactions. This is very different from radiation chemistry where the radiation levels are kept too low to
influence the chemistry. It is
the branch of chemistry concerned with the chemistry of radioisotopes, elements, and substances, the laws 
governing the physicochemical behavior of this radioactive matter, the chemistry of nuclear transformatio
ns, and the physicochemical processes that accompany these transformations. Radiochemistry, because of 
the topics, methods, and objects of its investigations, can be subdivided into general radio-

DETERMINATION OF RADIONUCLIDES

Radiation type
Radionuclide
Alpha Beta Gamma
Cesium 137   X X
Iodine 131   X X
Plutonium X X X
Radium X    
Strontium 90   X  
Tritium   X  
Thorium X   X
Uranium X   X
Radiometric and radiochemical methods as well as ICP/SFMS methods are used in ALS for the
determination of natural and artificial radionuclides in all the matrices from the Environment and in
food/feed samples. It can be said that we can determine all the common radionuclides from the
environment by the combination of the above methods. High Resolution Gamma-Ray Spectrometry
can be usually used without modification, samples with low concentrations of radionuclides are
concentrated (evaporation, separation on sorbents or by co-precipitation) prior
measurement. Radiochemical methods are modified according to the matrix composition .
Application of radiochemistry

By using especially sensitive methods for monitoring radioactive decay, it is possible to detect the presen
ce of single atoms of a radioisotope and to establish the fact of their decay-coworkers. Methods for the pr
oduction of nuclear fuel, the separation of Pu and fission products from the U irradiated in nuclear reactor
s, and the reprocessing of spent U from reactors have all been undergoing improvement. A number of imp
ortant technological problems related to nuclear fuel have been solved. The chemistry of artificial, especia

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lly transuranium, and natural, especially U, Th, and Pa, radioactive elements has undergone extensive dev
elopment, particularly in the chemistry of the complexes formed by these elements. The chemistry of new 
atomlike species—positronium, muonium, and mesic atoms—has also been established.Extraction and ch
romatography have acquired special importance in radiochemistry. 

Radiometry
What is Radiometry?
Radiometry is an instrument for detecting and measuring the intensity of radiant energy, by exposing
to sunlight a set of vanes blackened on one side and suspended on an axis in a vacuum and measuring
their speed of rotation (i.e., the mechanical energy into which the radiant energy has been
converted). It is used for physical measurement of a wide range of radiation from x-ray to radio wave.
Radiometric Instruments
Various types of instruments can be used for measuring radiometric quantities:
Optical power meters can be used for measuring a radiant flux, e.g. generated by a laser, and optical
energy meters for an energy. For highly divergent light, such devices are normally not usable. Here,
one may need to use an integrating sphere in conjunction with a suitable detector, e.g. a bolometer or
a detector. A photodiode can be used for measuring an irradiance, if it is calibrated for
the wavelength of some quasi-monochromatic light and its active area is small enough for the
required spatial resolution. The irradiance is the radiant flux divided by the active area
Radiometric Quantities
Various terms used in radiometry are not identical to those which are common
in  optics and laser technology. The following table also specifies such alternative terms
which are used particularly in optics:

 Radiometric  
Definition
Unit    Symbol  

The total optical power emitted from a source.

Radiant flux Φ
 

Irradiance E The flux per unit area striking a surface.

Intensity I The flux per unit solid angle from a source. (often confused

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 with irradiance or radiance)

The flux per unit solid angle, per unit projected area from
Radiance L
an extended source.

Luminous Flux Perceived power from a source weighted to the sensitivity

Φv
(Lumens) of the human eye

Radiant Exitance

In radiometry, another quantity exists which is less useful in practice but, of which, it could be useful
giving a definition. This quantity is radiant existence.

While irradiance concerns with the radiant power incident on a surface, radiant existence can be seen
as a “source quantity.” In fact, it is defined as the radiant flux emitted by a source per unit surface. In
this case, the surface is not that of the receiver but that of the emitter. As irradiance, its unit is W/m2.
Me=dΦdAMe=dΦdA

What Do Radiometric Units Reveal?

Radiometric units can be used to give a complete account of the power of sources such as
the sun, LEDs, light bulbs, and so on. This is vital since the sensitivity of any radiometric
measurement system could vary depending on the variations in all the components such as
optical components and the detector. The process of calibration enables the absolute
standard to be referred to derive calibrated radiometric units. Examples of radiometric
measurement applications can be found in quite a few research projects and industries.
Radiometric units allow us to make absolute statements about the power of sources ranging
from the sun to light bulbs or LEDs. 

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 Conclusion

Generally we understand from this all topic that for some instruments in the laboratory how to use it
and also who to deal with them. Additionally importance of this laboratory instruments and limit of
use them. To add one more we know how to calibrate with standard unit and for some instruments
what reason behind to work and break during some error as happen.

Reference

1. DeCusatis C (1997) Handbook of applied photometry. American Institute of Physics Press, New
York
2. Wolfe WL (1998) Introduction to radiometry: tutorial text in optical engineering, vol TT29. SPIE
Press, Bellingham
3. Parr AC, Datla RU, Gardner JL (2005) Introduction to optical radiometry. Elsevier Academic Press,
Amsterdam, 1 Jan 2005 Palladino P (2005) Manuale di Illuminazione. Tecniche nuove
4. 1. Cuatrecasas P, Wilchek M, Anfinsen CB. Selective enzyme purification by affinity
chromatography. Proc Natl Acad Sci U S A. 1968;61:636–43. [PMC free article] [PubMed] [Google
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5. 2. Porath J. From gel filtration to adsorptive size exclusion. J Protein Chem. 1997;16:463–
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