COMPLEMENT FIXATION.pptx

PG AND RESEARCH DEPARTMENT OF MICROBIOLOGY,SRI PARAMAKALYANI COLLEGE,ALWARKURICHI-627 412

1. SRI PARAMAKALYANI COLLEGE ( REACCREDITED WITH B GRADE WITH A CGPA OF 2.71 IN THE II CYCLE BY NACC AFFILIATED TO MANONMANIAM SUNDARANAR UNIVERSITY,THIRUNELVELI) ALWARKURICHI -627 412 TAMILNADU INDIA. POST GRADUATE & RESEARCH CENTER – DEPARTMENT OF MICROBIOLOGY (GOVERMENT AIDED) II SEM - CORE –IMMUNOLOGY UNIT – III COMPLEMENT FIXATION SUBMITTED TO THE GUIDE DR.S.VISWANATHAN,PH.D HEAD OF THE DEPARTMENT SRI PARAMAKALYANI COLLEGE ALWARKURICHI. SUBMITTED BY ESSAKIMUTHU .G REG.NO : 20211231516109 I M.SC, MICROBIOLOGY DATE: 30.04.2022

2. Definition Complement fixation is one of the most important and one of the classical techniques for determining antigen-antibody complexes present in the testing sample.

3. Principle When antigen and antibody interact with each other, they form a complex called antigen-antibody (Ag-Ab ) complex. The complex then interacts with complement protein and gets fixed with it. After fixing, the complement degrades or gets cleaved into two fragments i.e. smaller and larger fragments.

4. Positive test Antibody in sample + Antigen (added) + Complement    →     Ag-Ab Complex Fixed with Complement Complement fixed Ag-Ab + Indicator System     →     No change (No hemolysis)

5. Negative test Sample with no antibody + Antigen (added) + Complement    →     Free Complement Antigen (added) + Antibody in indicator system (On RBC)     →      Ag-Ab complex Ag-Ab complex + Complement     →     Fixed Complement System    →    Hemolysis

6. Complement Fixation Test   Requirements Samples such as serum or CSF (may or may not contain the specific antigens or antibodies of interest) Known complementary antigens based on the component desired to be detected.

7. Complement Proteins The native complement present in the sample is inactivated. Complement obtained from the serum of other organisms such as Guinea pig is added to the sample during the test.

8. I ndicator System Sheep erythrocytes or RBCs coated with antibodies(mainly derived from Rabbit serum) on the surface. These RBCs can also be called sensitized RBCs. 

9. Complement Fixation Test   Procedure A serum sample is taken. It is then heated at about 56 °C to remove the complement proteins already present in the sample. The serum is then adsorbed with washed sheep RBC. It prevents interference in the test by anti-RBC antibodies which are cross-reactive.

10. 4.Then the antigen and complement are added to the sample. 5.It is then subjected to incubation at a temperature of 37 °C for 30 minutes. It provides conditions and time for the formation of the Ag-Ab complex. 6.And the indicator system is then added and the sample is observed for change due to occurrence or non-occurrence of hemolysis.

11. Complement Fixation Test   Applications Wasserman’s test is one of the complement fixation tests for the detection of syphilis. It is an  antibody  detection test. It can also be used for the detection of bacterial diseases caused by Mycobacterium pneumoniae, Bordetella pertussis, etc. It can be used for the detection of viral infections, and fungal infections such as Histoplasmosis, Cryptococcosis, etc.

12. Complement Fixation Test   Advantages Interpretation of the result after the test is easier. It can be used for the detection of a very small number of antigen or antibody components in the sample. It can be used for the detection of a variety of infections. It has good sensitivity.

13. Complement Fixation Test   Limitations It is one of the old methods not used much in current practices. It is slower and more complex in comparison to many easier rapid detection tests being used currently. It is difficult to perform and arrange the reagents used for it. Although it is one of the sensitive tests, it has less sensitivity than tests such as  ELISA .

14. References Goldsby R.A., Kindt T.J., Osborne B.A., (1999) Kuby Immunology, 4th edition, W.H.Freeman & Co Ltd. Parija S.C., (2009), Textbook of Microbiology and Immunology, 2nd edition, Elsevier, a division of Reed Elsevier India Private Limited Jeffrey K., Complement Fixation Test  Introductory Immunology (Second Edition) , 2019Miller, V. B. (1930). Tests for Syphilis: An Explanation of the Wasserman Test. The American Journal of Nursing, 30(6), 707–712.