Jimma University

Institute of health
Faculty of health sciences
School of medical laboratory sciences
Immunology& Molecular Biology course team

Immunology and Serology

course
Audiences: 3rd year regular Medical Labs
Instructor :Mr.Dejene G(MSC, PhD candidate)
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 Course syllabus
Week 8 Immunological and serological techniques 1hrs
o Overview of immunological and serological techniques
o Laboratory materials & equipment for immunologic and serologic
tests
o Laboratory animal handling and management
o Quality control in Immunology and Serology

Laboratory visit on Instrument in Serology laboratory 3hrs

Week 9 o Antigen-antibody reaction 4hrs

o Overview of antibody reaction
o Primary binding test
o Immunofluorescence, ELISA, Radioimmunoassay

Week 10 o Secondary binding tests 4hrs

o Precipitation Reactions, Agglutination, Complement fixation tests
o Tertiary binding tests

Week 11 o Flow Cytometer 4rs

o Methods of Monoclonal Antibody Productionion
Laboratory visit on Instrument in Serology laboratory 6rs
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 Course syllabus…
Week 12 o Specimen collection and preparation for serological tests 3hrs
o Safety, specimen collection, preparation, preservation and
Shipment of serological specimens.
o Dilution (Serial dilutions and Determination of end point
and titer) and Complement inactivation

Laboratory demonstration and practice on Serum 6hrs

preparation, Serum dilution and pipetting
Week 13 o Common Serologic Tests for Bacterial and Parasitic disease 4hrs
• Serological diagnosis for syphilis
• Syphilis (Characteristics of the Organism, Mode of
Transmission, Stages of the Disease, Congenital
Syphilis, Nature of the Immune Response)
• Serological diagnosis of syphilis
Laboratory visit and demonstration on Immunology laboratory 6hrs
equipment(ELISA, flow cytometry)

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 Course syllabus…
Week 14 • Agglutination test for febrile diseases 4hrs
o Serological test for Salmonella infection
o Serological test for Rickettsial infections
o Serology of Streptolysin O (SLO) and Anti-
streptolysin O (ASO)

Laboratory demonstration and practice on ASO,RPR and 6hrs

VDRL test for syphilis qualitative and quantitative
methods
Week 15 o Serological tests for Helicobacter pylori 4hrs
o Serology test for Malaria
o Serological tests for Leishmaniasis
o Serological tests Toxoplasmosis
Laboratory practice on selected bacterial and parasitic 6hrs
diseases
Demonstration on ELISA

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 Course syllabus…
Week • Common Serologic Tests for Viral Infections: 4hrs
16 o Serological tests for HIV
 Characteristics, Composition, Structural Genes,
Replication and Immunology of HIV
 Common HIV Antibody Tests
 Current HIV test algorithm
 Common HIV Antigen Tests
 Western blot
 Molecular technique (Viral load)

Laboratory practice on serological diagnosis of HIV and other 6hrs

viral diseases using rapid test kits and ELISA

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 Course syllabus…
Week 17 • Serology of Hepatitis Viruses 4hrs
• Serology of Infectious mononucleosis
• Serology of Dengue viruses
Laboratory practice on serological diagnosis of HIV and 6hrs
other viral diseases using rapid test kits and ELISA
Week 18 • Serologic test for Autoimmune disease 4hrs
o Diagnosis of systemic lupus erythematosus
o Serology of Rheumatoid Factor
o Acute-phase reactants/ Acute-phase Proteins (C-
reactive protein)
o Serology of Troponin I

Laboratory demonstration and practice on SLE, 4hrs

Rheumatoid Factor and CRP tests

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 Course syllabus …
Week 19 • Serology of Human Chorionic Gonadotrophin 4hrs
Hormone (HCG)
o Urine pregnancy tests, factors that affect urine
pregnancy test,
o Urine specimen collection
o Methods of determining HCG
Laboratory practice on Rheumatoid Factor and Urine 6hrs
pregnancy tests
Week 20 Final examination

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 Chapter 14

Immunological and serological techniques

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 Outlines
Overview of immunological and serological
techniques

Laboratory materials & equipment for

immunologic and serologic tests

Laboratory animal handling and management

Quality control in Immunology and Serology

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Overview of immunological and
serological techniques

What is immunology?

What is serology?

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 What is Immunology?
The study of:
o The tissues, organs and cells related to the immune
system

o The immune system’s response to a foreign body

or antigen which includes the production of
antibodies

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 What is Immunology?...

The study of:

o The immune system in response to allergens

o Autoimmune diseases

o The system in response to cancer cells

o Immunotherapy and

o Diseases or infections of the immune system

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 What is Serology?
 Serology is the scientific study of blood serum

 Serum is a part of blood

 It is usually formed by allowing blood to clot

 The process of clotting removes the clotting

factors and whole cells from the blood leaving
behind a yellow coloured liquid
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 These serum contains
 Antibodies
 Antigens
 Microorganisms
 Hormones
 Electrolytes and other proteins
 2-4 are hemolysed serum:not used
Normal serum
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 Serology, in a wider context basically
deals with
 The quantitative and qualitative analysis of these
various components
 However, serology is known for the qualitative
detection or quantitative analysis of antibodies or
antigens concerning infection or disease diagnosis
o In practice, the term usually refers to the diagnostic identification of
antibodies in the serum
 Such antibodies are typically formed in response to an infection
(against a given microorganism)

o Against other foreign proteins (in response, for example, to a

mismatched blood transfusion), or to one's own proteins (in instances
of autoimmune disease)

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 Immunology versus Serology
Immunology is the study of the immune
system and its functions and disorders

Serology is the study of serum

Both are related and connected to each other

and contribute to a better understanding of
disease and infection in the body
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 Immunology versus Serology…
The connections between both is reactions
found in immunology form the basis of
serology or serological techniques

In other words, serology can also be

considered a branch of immunology with it
focusing on the diagnostic values of the
immune system

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Immunology is concerned about INVIVO
antigen antibody reaction(inside the body)

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Serology is concerned about Invitro antigen
antibody reaction(outside the body)
Why Invitro antigen antibody reaction?
Assays detect presence of antigen or antibody
1. Diagnosis of infectious and non infectious
diseases
 Monitoring humoral immune response
 Monitoring therapeutics
 Analysis of interesting molecules

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1. Diagnosis of infectious and non infectious diseases…

 known antigen preparations are used to detect

circulating antibodies in patient's serum as
evidence of a current or previous infection
with that agent OR

 known antibodies are used to detect antigens

associated with an infectious agent directly in
body fluids
2. Identification of unknown cultures:
known antibodies are used to detect their
homologous antigens in cultures

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 Methods of In vitro Ag-Ab rxn
1. Precipitation reactions
(Ag is soluble) precipitation

2. Agglutination reactions
(Ag is particles) clumping

3. Complement fixation reactions.

Lysis
4. Labelling methods/immunoassys

 Immuno-fluorescence reactions
 ELISA/EIA also Chemiluminescence
 RIA
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 Immunoblotting
 Methods of In vitro Ag-Ab rxn..
 Almost all of these techniques are based on

o Reactions achieved in the context of specificity

between components of the immune system, i.e.
antibodies, and antigens
 The fine specificity of antigen-antibody interactions has
led to
o The development of a variety of immunologic assays
o Can be used to detect the presence of either antibody
or antigen and antigen antibody complex

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 Methods of In vitro Ag-Ab rxn…
 Immunoassays have played vital roles in:
o Diagnosing diseases
o Monitoring the level of the humoral immune
response
o Identifying molecules of biological or medical
interest
 These assays differ in their speed and sensitivity
 Some are strictly qualitative, others are quantitative

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 Application of serological tests
For diagnostic purposes when an infection is
suspected

In rheumatic illnesses, and

In many other situations, such as checking an

individual's blood type

Human Leukocyte Antigen (HLA) Typing

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 Application of serological tests…
 In the field of forensics aiding in crime solving

 In epidemiology

 To diagnose patients with certain immune

deficiencies associated with the lack of
antibodies

 In such cases, tests for antibodies will be

consistently negative

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 Applications of serology and immunology
Antigen tests often enable an early diagnosis
or presumptive diagnosis of an infectious
disease through:-

Identification of a pathogen that has been isolated

by culture

Identification of pathogens in different samples of

the patients, etc

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 Applications of serology and immunology…

Antibody tests are used mainly:-

To diagnose a microbial disease when the pathogen

or microbial antigen is not present in routine
specimen or

If present is not easily isolated and identified by

other available techniques.

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 Applications of serology and
immunology…
Antibody tests are used mainly:-

To screen donor blood for different infectious

diseases

To monitor the effectiveness of a given

treatment by measuring antibody liter

To diagnose autoimmune disorders, etc.

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 Summary
 Immunology is the study of the immune system majorly in a
state of in vivo
 Serology majorly refers to the in vitro study of blood serum
rather than the study of it is in vivo nature
 Serology is a comparatively smaller discipline compared to
immunology
 Immunology has a relatively wider scope than serology
 Serology is the science of measuring antibody or antigen in
body fluids

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 Laboratory materials & equipment
for immunologic and serologic tests

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 List of materials and equipment
Centrifuges, thermal water baths
ELISA reader
ELISPOT automated reader (Immunospot)
UV/VIS spectrophotometer
Modern visible light and fluorescent
microscopes

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 List of materials and equipment…
Glass wares
o Test tubes

o Glass slides, cards

o Serological pipette with a different size

Plastic wares
Constant temperature devices
 Rotating machines
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 List of materials and equipment
Reagents
o House made
o commercial
Specimens
o Whole blood
o serum, plasma, urine, CSF
o peritonial,pericardial,plurial and etc

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 List of materials and equipment…
Glass and Plastic wares
 Test tubes of different sizes
 Capillary tubes
 Glass/plastic/cartoon slides
 Glass slides with wells (flocculation slides)
 Cylinders, beakers, volumetric flasks of different
volume
 Micro titration plates
 Serological pipette with a size of 10ml, 5ml, 2ml 1ml
and less (0.2 ml).
 Pipettors and pipette tips (plastic)
 Reagent bottles of different size
 Reagent dispensers/droppers
 Dropping bottles Mr Dejene G(Medical Immunologist)
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 List of materials and equipment. . .

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 Glass and Plastic wares
 Dirty glassware easily affects serological
tests
 After using all the glassware (test tube,
beaker, pipette, etc) they should be soaked in
detergent for several hours and rinsed several
times in tap water
 Finally, allow drying by placing in a dry oven
or dust free place. Test tubes and pipettes
should not be scratched or broken, which will
interfere with the reading of a test
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 Constant temperature devices
Incubators –usually 37oc
Water baths-usually 56oc
Refrigerators-usually 2-8oc
Deep Freezers-usually -20,-70 and -80oc
o All are electrically operated and have thermostat that
hold the temperature within the required limits

o These should be checked prior to use by a thermometer

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 Rotating machines
to facilitate antigen antibody reactions
have a flat plate, which rotate at a prescribed
rate of speed
A knob located on the front of the machine
controls the number of revolutions per minute.
 rotate in different directions:
▫ Circular ,Rectangular, back and forth

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 Materials necessary for basic serologic tests…
Capillary tubes
Yellow tips

Serological card slides

Serological reagents Flocculation slides

Serological glass slide

Multichannel
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pipette Mr Dejene G(Medical Immunologist) 39
Micro titration plate well Dropping botteles
 ELISA washer (microplate washer)

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 Microtiter plate/Microplate

• It is also called microwell or multiwall, which is a flat plate

with multiple "wells" used as small test tubes. The
microplate has become a standard tool in analytical
research and clinical diagnostic
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Immunologist) 41
 Microplate Readers

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 Serofuge

A serofuge is a small centrifuge used in

blood banking and serology to spin
serological tubes.

A serofuge is a small centrifuge

specifically designed to separate serum
from cells and to complete blood cell
washing procedures. Like a centrifuge,
a serofuge has holes to insert the
centrifuge tubes in, a lid to protect
both the samples and experimenter in
case of malfunction, and a way to set
the centrifuge speed and way to turn
the centrifuge on and off.
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Laboratory animal handling and management

SOPS guides are required

Training and supervision required
Environment must be controlled
Disease or pathogen free Lab rodents
o Don't posses antibody show past infections
o Available from commercial vendors
Disease free primates

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 Purpose of lab animals
In vivo administration of antibodies to
experimental animals
o To manipulate the immune system
To synthesize known antibodies by injecting
known antigens

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 Common Lab animals
• Includes
Rats
Mice
Rabbits and guinea pigs
Hamsters
Non human primates
Amphibians, fowl, sheep and pigs
Mice and rats make up approximately 95% of all
laboratory animals, with mice the most commonly
used animal in biomedical research

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 Requirements considered
Housing
Bedding
Feeding
Water
Sanitations
Vermin control

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 Quality control in Serology Lab
We consider:
o Preanalytical

o Analytical

o Post analytical

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 Thank you

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