The document discusses antigen-antibody reactions and their uses and characteristics. It describes how antigen-antibody interactions form the basis of immunity against infectious diseases and can also cause hypersensitivity reactions or tissue damage. It outlines several serological tests used to detect antibodies or antigens in the laboratory, including agglutination reactions, precipitation reactions, complement fixation tests, neutralization tests, and immunoassays. It provides details on the mechanisms and applications of specific tests like slide agglutination, tube agglutination, latex agglutination, and complement-dependent cytotoxicity tests.
The document discusses antigen-antibody reactions and their uses and characteristics. It describes how antigen-antibody interactions form the basis of immunity against infectious diseases and can also cause hypersensitivity reactions or tissue damage. It outlines several serological tests used to detect antibodies or antigens in the laboratory, including agglutination reactions, precipitation reactions, complement fixation tests, neutralization tests, and immunoassays. It provides details on the mechanisms and applications of specific tests like slide agglutination, tube agglutination, latex agglutination, and complement-dependent cytotoxicity tests.
The document discusses antigen-antibody reactions and their uses and characteristics. It describes how antigen-antibody interactions form the basis of immunity against infectious diseases and can also cause hypersensitivity reactions or tissue damage. It outlines several serological tests used to detect antibodies or antigens in the laboratory, including agglutination reactions, precipitation reactions, complement fixation tests, neutralization tests, and immunoassays. It provides details on the mechanisms and applications of specific tests like slide agglutination, tube agglutination, latex agglutination, and complement-dependent cytotoxicity tests.
The document discusses antigen-antibody reactions and their uses and characteristics. It describes how antigen-antibody interactions form the basis of immunity against infectious diseases and can also cause hypersensitivity reactions or tissue damage. It outlines several serological tests used to detect antibodies or antigens in the laboratory, including agglutination reactions, precipitation reactions, complement fixation tests, neutralization tests, and immunoassays. It provides details on the mechanisms and applications of specific tests like slide agglutination, tube agglutination, latex agglutination, and complement-dependent cytotoxicity tests.
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ANTIGEN - ANTIBODY
REACTIONS
Dr.Gomathi M.D., Prof. Dept of Microbiology SRM MCH & RC Ag-Ab reactions - USES (i) In vivo (In the body)
• Protection against microbes and their products.
• Forms the basis of Ab mediated immunity in infectious diseases.
• Forms the basis of tissue injury in some types of hypersensitivity &
auto-immune diseases.
(ii) In vitro (in the lab-serological tests)
• To detect the presence of Ab or Ag.
• Vital role in diagnosing diseases.
• Monitoring the level of the humoral immune response
• Identify molecules of biological / Medical interest.
Ag-Ab interaction 1.Primary stage : No visible effect. Reaction is rapid. Reaction is reversible. Occurs by ionic bonds, H2 bonding. Free & bound Ag/Ab can be estimated by physical or chemical methods 2.Secondary stage: Leads to Precipitation, Agglutination, lysis of cells, Neutralisation of toxins , Complement fixation test, Immobilisation of microbes, enhancement of phagocytosis – demonstratable. 3.Tertiary reaction: some Ag-Ab reactions leads to chain reactions resulting in neutralisation or destruction of injurious antigens or to tissue damage. General characters of Ag-Ab reactions • Highly specific. (In some cases Ab elicited by one Ag can cross react with an unrelated Ag due to antigenic similarity) • Both Ag & Ab participate in lock & key arrangement. • No Denaturation of Ag/Ab during the reaction. • Surface antigens are most relevant during combination. • Entire molecules react (Not fragments). • Combination is firm but reversible, firmness depends upon the affinity (attraction) & avidity (Strength of bond). • There is no chemical alteration in either the Ab or Ag • Combine in varying proportions. Ag – Ab Measurement • Measurement of Ag / Ab is in units / titer
• Titer is the reciprocal of the highest
dilution of the serum which shows and observable reaction Parameters of serological tests Sensitivity:
• Ability to identify correctly all those who have the disease.
• True positives • False negatives – low / nil.
Specificity:
• Ability to identify correctly all those who do not have the
• More sensitive than precipitation because of the
direct nature of antigen-antibody interaction AGGLUTINATION Visible clumping of particles Better with IgM TYPES OF AGGLUTINATION • Slide agglutination • Tube agglutination • Antiglobulin (Coomb’s) test • Heterophile agglutination test • Passive agglutination (indirect) Latex agglutination, Haemagglutination, Co-agglutination SLIDE AGGLUTINATION TEST
Uses: To identify bacterial isolates within 30-60 seconds Method of Blood grouping & cross matching Rapid & convenient Blocking antibodies
• Occasionally antibodies are
formed that will react with the antigenic determinants on a cell but will not cause agglutination. Ex Anti Rh antibodies and anti brucella antibodies TUBE AGGLUTINATION TEST Uses • Quantitative methods for detection of Abs • Serological diagnosis of Enteric fever - Widal test Brucellosis - Brucella agglutination test Rickettsial fevers- Weil Felix test Infectious mononucleosis- Paul Bunnell test Primary atypical pneumonia - Cold agglutination test Streptococcal MG agglutination test
U U U U UU U U U HETEROPHILE AGGLUTINATION TEST • Due to common Ag sharing in unrelated species Weil Felix test Paul Bunnell test Cold agglutination test Streptococcal MG agglutination test PASSIVE AGGLUTINATION TEST • Precipitation reaction converted into agglutination
• Soluble Ag is attached to the surface of
carrier particles
• Latex, bentonite & RBC(s)
REVERSE PASSIVE AGGLUTINATION • Ab is absorbed to carrier particle
• For estimation of Ag
• Eg: Detection of HBsAg using RPHA
LATEX AGGLUTINATION TEST • Latex particles can be coated with Ag/Ab • Eg: Anti Streptolysin O C Reactive Protein Rheumatoid Factor LATEX AGGLUTINATION TEST HAEMAGGLUTINATION TEST • RBC(s) senstised with Ag are used for detection of Abs • Eg: Rose-Waaler test CO-AGGLUTINATION TEST USE • For detection of bacterial Ag in serum, urine & CSF • Neisseriae, Str.B, C, D, F, G & N & H.influenzae Ags A-F NEUTRALISATION TEST Principle: • When antitoxin combines with a toxin, the biological effect of toxin gets neutralised • Since toxin is an Ag in solution, it is precipitated • Eg: Viral neutralisation tests Neutralisation of bacterial toxins VIRAL NEUTRALISATION TESTS
Test serum + Normal serum +
viral suspension viral suspension
Embryonated egg / Embryonated egg /
tissue culture tissue culture
No evidence of infection Evidence of infection
NEUTRALISATION OF BACTERIAL TOXINS • In vivo Schick test: To demonstrate immunity to Diphtheria intradermally • In vitro Nagler reaction: To demonstrate lecithinase production by Clostridium perfringens ASO (antistreptolysin) test NAGLER REACTION
Lecithinase Antiserum COMPLEMENT FIXATION TEST Principle: • When complement binds to an Ag – Ab complex it becomes “fixed” and “used up”
• CFT involves two steps and five reagents
(Ag ,Ab , compliment, sheep cells and amboceptor CFT – Controls • Ag & serum control to ensure that they are not anti complementary • Complement control to ensure that desired amount of complement is added • Cell control to see that they do not undergo lysis in the options of the complement COMPLEMENT FIXATION TEST
Uses: • Retrospective diagnosis of viral, Fungal , Rickettsial, Chlamydial, Protozoan infections • Wassermann’s reaction for syphilis COMPLEMENT DEPENDANT TESTS • Immobilisation test ( eg. TPI) Motile treponemes treated with patient’s serum in presence of complement
• Immune adherence (eg.V.cholerae)
Ag-Ab complex adhere to particulate materials (RBC/ platelet/macrophage) in presence of complement
Contd… COMPLEMENT DEPENDANT TESTS Contd… • Cytolytic or cytocidal reaction Live bacteria when mixed with specific Ab in presence of complement will produce bacteriolysis Eg: Vibrio cholerae Indirect CFT • Certain sera do not fix g.pig complement • Setup test in duplicate • After first step standard antiserum known to fix complement is added to one set • If test serum is positive , antigen would have been used up. There fore standard serum will not fix complement • Therefore hemolysis indicates a positive result in the indirect CFT Conglutinating complement absorption test
