detection of Rheumatoid factor by using LatexAgglutination

1. detection of Rheumatoid factor by using LatexAgglutination Dr Laila Hammed Damanhouri

2. Agglutination test • It is one of important laboratory method to detect antigen antibody reaction. • It provides flexible and useful method for semi quantitating of either antigen or antibody concentration. • The reaction occurs between insoluble antigen and appropriate antibody. • The reaction will results in forming aggregate or agglutinate.

3. Phase one Antibody reacts with single antigenic determinants on or close to particle surface. It is a rapid reaction. Stages of agglutination reaction

4. A single antibody molecule binds to antigenic determinants on adjacent particles. The visible reaction occur under appropriate conditions and over time, particles remain connected and interconnected by antibody bridge. Secondary phase

5. To test patient’s sera (contain antibody) against large antigen. Direct agglutination can be used to determine antibody titer. Types of agglutination reaction1. Direct agglutination.

6. serum is mixed with latex spheres (inert substance) with the soluble antigens attached. Antibodies will then cause visible agglutination of the latex spheres with the soluble antigens attached. Indirect agglutination

7. the red blood cell are coated with soluble . then incubated with patient serum (contain AB against Ag. the interaction between AB in the patients sera and antigen on the surface of red blood cell resulting on agglutination of the red blood cell If the reaction not occur the red blood cell will form as a button-shaped deposit at the bottom of reaction vessel. Indirect hemagglutination

8. advantages of agglutination methods • ease of performance. • speed of performance, usually requiring few minutes. • high degree of sensitivity.

9. Disadvantages of agglutination methods • the reaction are only semiquantitative. • the occurrence of the prozone phenomenon, in which agglutination is inhibited by extreme antibody excess as a result of poor lattic formation.

10. prozone • Absence of agglutination at higher antibody concentration. • It is due to many factors including • Presence of blocking antibodies at low titers • Inaccessible antigenic determinants • Weak avidity • Poor lattice formation. • The problem can be avoided by use of standard serial dilution.

11. Application of agglutination test • several antibodies can be detected by this method such as Rheumatoid factor.

12. his test is done to diagnosed Rheumatoid arthritis, which is one of important autoimmune disease. RF is an antibody ( IgM or IgG classes) bind to the Fc portion of other IgG molecules, and form IgG-anti-IgG complexes in the circulation or joint fluid. Rheumatoid Factor (RF)

13. Rheumatoid factor (RF) • RFs are detected in serum in up to 80% of adult patients with RA. • RFs are not specific for RA and occur in other autoimmune disease, in chronic infectious diseases, such as infective endocarditis, tuberculosis, and hepatitis B. • usually at low titer, in up to 20% of overtly normal elderly individuals

14. General Feature of Rheumatoid arthritis • RA is a systemic chronic inflammatory disease of unknown etiology. • is characterized by polyarthritis which may be progressive and permanently deforming and by extra-articular manifestations(rheumatoid nodules, pericarditis, and arteritis). • Adult RA is commonly associated with rheumatoid factors.

15. Symptoms first begin in the small joints of the fingers, wrists and feet, with warm, swollen and tender joints that are painful and difficult to move. Joints of both sides of the body (symmetrical) are typically affected. People with RA often experience fatigue, loss of appetite and low-grade fever. There is often stiffness in the morning that lasts for several hours or more. Nodules may form under the skin, often over the bony areas exposed to pressure (such as the elbows). Over time, damage to the cartilage and bone of the joints may lead to joint deformities. Symptoms of Rheumatoid Arthritis

16. diagnosis of rheumatoid arthritis • medical history and physical examination, looking for distribution of joints affected, joint swelling, warmth and range of motion, as well as the presence of nodules under the skin. • Imaging studies such as X-rays, sonograms or magnetic resonance imaging may be used to detect the degree of joint involvement or joint damage. • A blood test can indicate the presence of an rheumatoid factor, which is found in 80 percent of people with RA.

17. X ray

18. Methods used to detect RF • Latex agglutination method. mixes the blood being tested with (latex) beads that are covered with human antibodies. If rheumatoid factor (RF) is present, the latex beads clump (agglutinate). • Haemagglutination test. mixes the blood being tested with a sheep's red blood cells that have been covered with rabbit antibodies. If RF is present, the red blood cells agglutinate. • Nephelometry test Using an automated machine based on laser light scattered.

19. Latex agglutination test

20. Interpretation of the test • Agglutination test is positive. Do titration and determine the end of titration. • Normal range differ from lab to another, but in most lab titration of >1:20 consider positive. • Positive test in 80% of Rheumatoid Arthritis. • It also positive in other autoimmune disease. • Positive in viral hepatitis. • Positive in TB.

21. Factors interfere with positive result • Hyperlipedemia. Blood that is very high in fats. • Age. About 5% to 10% of people over age 65 have an elevated RF level.