Exp Ⅱ Agglutination reaction and Antibody preparation (2)

Test 1 Agglutination reaction

1.Principle: Tube agglutination reaction Ag-Ab reaction Agglutination reaction Slide agglutination reaction QualitativeSemi-Quantitative Test 1 Agglutination reaction

1.Principle: Test 1 Agglutination reaction 1) Slide agglutination reaction: Known Ag + Unknown Ab Agglutination turbid ? 2) Tube agglutination reaction: Quantitative Ag + Serially diluted Ab Different degree of agglutination YES NO Determine the titer of Ab

1.Principle: What is the titer of Ab? The titer is customarily reported as the reciprocal of the highest dilution of Ab that causes an obvious agglutination. Test 1 Agglutination reaction

2. Experiments Ⅰ Tube agglutination test Ⅱ Slide agglutination test Test 1 Agglutination reaction

Ⅰ Tube agglutination test 1. Material: Inactivated typhiod bacteria (7×10 8 /ml), anti-serum 2. Method: :201:40 1:801:1601: Discard N.S Anti-H Serum Control 0.5ml ① ② ③ ④ Inactivated typhiod bacteria 0.5ml0.5ml0.5ml0.5ml0.5ml0.5ml Dilution ratio of Ab Shake 56 ℃, 1~2h

3. Attentions  Sign the tube  Use the dropper tube correctly  Mix thoroughly  T: 56 ℃  Record the time  Take the tube carefully Ⅰ Tube agglutination test

4. Anticipated results + Very small agglutination turbid -No agglutinationSame as control ++ Small and depositing partly Obvious turbid +++ Large and depositing mostly Slightly turbid ++++ Large and depositing Clear Degree of agglutination SedimentSupernantant Ⅰ Tube agglutination test

Sediment Agglutination degree The highest dilution ratio Titer of the anti-serum

Ⅱ Slide agglutination test 1. Material: Staphylococci Inoculation loop 2. Method: 12 Anti serum1 a. Slide b. Bacteria c. Reaction Mix bacteria with the serum d. observation Anti serum2

3. Anticipated results:+- Ⅱ Slide agglutination test

4. Attentions:  Sterile operation Ⅱ Slide agglutination test

Test 2 Secondary immunization

2. Methods: Uncoagulated SRBC Add 3ml N.S and mix 2000rpm, 5min Discard the supernant Make 20% 、 25% 、 30% 、 40% SRBC solution Take 0.1ml to immunize the mice subcutaneously Add 3ml N.S and mix