Storage method of bacterial liquid for counteracting toxic substances in animal vaccine effectiveness test
A preservation method and technology of bacterial liquid, which is applied in the direction of bacteria and microorganism preservation, can solve the problems of large fluctuations in the number of challenged bacteria and heavy workload, saving financial resources, manpower and material resources, long storage time, and ensuring stability and accuracy effects
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Embodiment 1
[0024] (1) Streak inoculation of the bacterial strains used for attacking the virus on the blood Martin agar medium, and culture at 37° C. for 24 hours;
[0025] (2) Pick typical colonies to inoculate serum-containing Martin broth medium, culture at 37°C for 16 hours, and count viable bacteria. The result of viable bacteria count is 5.32×10 9 CFU / mL.
[0026] (3) Then mix the bacterial solution with a vortex shaker, and distribute it into sterilized cryopreservation tubes, 1 mL per tube, numbered A1~A10, B1~B20, among which A1~A10 should be stored at 4°C, and B1~B10 should be stored at 4°C. Store at -80°C.
[0027] 2. Survival rate test
Embodiment 2
[0029] ① On the second day, take A1~A5 and B1~B5 respectively for multiple viable counts, and the counting results are shown in Table 1:
[0030] Table 1 The situation of live bacteria after the bacteria used for challenge were stored at 4°C and -80°C for 1 day
[0031] group
Viable Bacteria (CFU / mL)
group
Viable Bacteria (CFU / mL)
A1
4.35×10 9
B1
5.31×10 9
A2
4.44×10 9
B2
5.32×10 9
A3
4.51×10 9
B3
5.30×10 9
A4
4.39×10 9
B4
5.29×10 9
A5
4.28×10 9
B5
5.32×10 9
Average number of live bacteria
4.394×10 9
Average number of live bacteria
5.308×10 9
82.59％
99.77％
[0032] ② On the third day, take A6~A10 and B6~B10 respectively for multiple viable counts, and the counting results are shown in Table 2:
[0033] Table 2 The situation of live bacteria after the bacteria used for challenge were stored at 4...
Embodiment 3
[0041] In order to verify the toxicity of the bacterial solution preserved at -80°C, an animal experiment was carried out: SPF chickens were divided into 4 groups, 10 in each group, and the neck was injected subcutaneously.
[0042] Group 1: Newly cultivated bacterial liquid, adjust the concentration of the bacterial liquid, the number of challenged bacteria is 3 CFU / body, 10 / 10 died
[0043] Group 2: Store at -80°C for 30 days, adjust the concentration of the bacteria solution, the number of challenge bacteria is 3CFU / body, and 10 / 10 die
[0044] Group 3: Store at -80°C for 35 days, adjust the concentration of the bacteria solution, the number of challenge bacteria is 3CFU / body, and 10 / 10 die
[0045] Group 4: sterile saline, 10 / 10 healthy and alive.
[0046] The test results showed that the bacterial liquid was stored at -80°C for 35 days without affecting the virulence of the bacterial liquid, and could be used in the challenge protection test after vaccine immunization. ...
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