Preparation method of hemagglutination inhibition antigen of avibacterium paragallinarum and application of antigen
A technology of hemagglutination inhibition and poultry, which is applied in the field of preparation of hemagglutination inhibitory antigens, can solve the problems of low antigen titer, loss of hemagglutination activity, economic loss, etc., and achieve good precipitation and aggregation, excellent hemagglutination activity, highly operable effect
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Embodiment 1
[0050] Example 1 Preparation and Preservation of the Hemagglutination Inhibiting Antigen of Avibacterium paragallinarum
[0051] 1. Take Avibacterium paragallinarum (provided by the Animal Husbandry and Veterinary Laboratory of Beijing Academy of Agriculture and Forestry Sciences), mark it on a TSA plate, and place it in a candle jar at 37°C for 16h-28h, until the colonies grow out. Pick colonies from the purely cultured plate, inoculate 100mL TSB medium, incubate at 37°C for 10h-16h, harvest the culture when OD600≥0.9, centrifuge the culture at 8000×g for 10min, wash the precipitate with PBS for 3 times, Then suspend with 20mL-30mL PBS solution to obtain PBS suspension of Avibacterium paragallinarum.
[0052] 2. Take Staphylococcus suis (provided by the Institute of Animal Husbandry and Veterinary Medicine, Beijing Academy of Agriculture and Forestry Sciences), streak culture on ordinary broth agar medium, inoculate into LB liquid medium for overnight culture after the coloni...
Embodiment 2
[0056] Example 2 Preparation of 10% Aldehydelated Red Blood Cells
[0057] 1) Preparation of 10% fresh erythrocytes. The erythrocytes collected from healthy non-immune roosters were mixed with Alfred’s solution at a volume ratio of 1:1. After centrifugation at 500×g, the precipitate was washed with normal saline for 5 times. 10 times the volume of red blood cells, each centrifugation × 500g, centrifuged for 10 minutes, and the precipitated red blood cells were made into a 10% red blood cell suspension with a volume ratio of phosphate buffer (PB solution).
[0058] 2) Prepare glutaraldehyde solution (50%) according to the ratio of 5% of the above red blood cell suspension volume. First, dilute the glutaraldehyde stock solution from 50% to a final concentration of 5% glutaraldehyde with PBS, and place on a magnetic Put it on the stirrer, add it into the red blood cell suspension while stirring (if the volume of the red blood cell suspension is 100 ml, then take 5 ml of 50% gluta...
Embodiment 3
[0064] Embodiment 3 Hemagglutination (HA) test and hemagglutination inhibition (HI) test of Avian bacilli paragallinarum
[0065] 1. Hemagglutination (HA) test
[0066] The operation is best carried out at room temperature within the range of 20°C to 25°C. If the room temperature is not within this range, it can also be placed in a 37°C incubator. Take care to cover with a lid or parafilm to prevent the liquid from evaporating.
[0067] 1) HA test operation method: use a 96-well V-shaped microplate for the test, and the total reaction volume is 75 μl. According to Table 1, use bovine serum albumin PBS (abbreviated as PBSS, its configuration method is: take 0.1g bovine serum albumin V, dissolve it in 100ml PBS, filter and sterilize, put 2~8 ℃ for preservation.) Example 1 The antigen prepared in the method was diluted 5-fold first, then serially diluted 2-fold, then added 1% aldehylated red blood cells, mixed gently, and left at room temperature (15-25°C) for 40-60 minutes or ...
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