SN/T 1488-2004Protocol of haemagglutination-inhibition test for viral haemorrhagic disease of rabbit (English PDF)

3.1 Instrument

Ordinary balance, analytical balance, common centrifuge, 96-well V-type blood coagulation plate, 25 µL adjustable pipette, incubator, micro-oscillator, etc.

3.2 Preparation of suspicious rabbits and normal rabbit liver suspension

The liver was cut separately, and the physiological saline was added 1.10, homogenized, centrifuged at 4000 r/min for 30 min, and the supernatant was taken for hemagglutination.

test.

3.3 Saline

A physiological saline solution of 0.85% concentration was prepared.

See Appendix A for formulation.

3.5 Preparation of 1% human "O" type red blood cell suspension

Take fresh human "O" type red blood cells in a solution (preserved at 4 ° C for one week). Take human "O" type red blood cells at a time of 20 times the washed red blood cells were mixed with saline, centrifuged at 2000 r/min for 15 min, the supernatant was discarded, and the washing was repeated four times. The washed red blood cells were mixed with physiological saline.

The 1% red blood cell suspension was set at 4 ° C for use and used up within 72 hours.

3.6 Antigen, positive serum

It is supplied by the designated unit and used according to the instructions.

3.7 Treatment of tested serum

Blood samples were taken aseptically, serum was separated, and serum was inactivated at 56 ° C for 30 min.

4.1.1 Experimental steps

4.1.1.1 Each sample was made in the same 2 rows, and 25 µL of physiological saline was added from the first well to the 11th well.

4.1.1.2 The first and second wells were added with suspicious rabbit liver tissue suspension 25 µL.

4.1.1.3 Start from the second well, dilute to the 10th well with a 25µL pipette, and discard 25µL in the last well (the virus is diluted) the degree is 10, 20, 40, 80, etc.).

4.1.1.4 The 12th hole is a normal rabbit liver suspension control.

4.1.1.5 In the second row, add 25 µL of positive serum to each well, shake well on a micro-oscillator, and place in a 37 ° C incubator for 10 min.

4.1.1.6 Add 25 µL of 1% human "O" type red blood cells to each well, shake immediately on the micro-vibrator, and set the refrigerator for 4 minutes at 45 °C.