Animal brucella antibody colloidal gold test paper film detection reagent kit
A technology of colloidal gold test paper and Brucella, which is applied in the field of animal epidemic diagnosis, can solve the problems of time-consuming, laborious, and inability to distinguish the pathogenic types of Brucellosis, and achieve the effect of rapid detection
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Embodiment 1
[0032] 1. Extraction and identification of animal Brucella LPS
[0033] The LPS of Brucella bovis 544A and Brucella sheep 16M were extracted by hot phenol method and cold phenol method respectively, and purified by Sephadex G-50 chromatography column, and the purity and immunological characteristics of the purified LPS were tested. Identification. The results showed that the content of LPS was 1.21mg / mL-2.31mg / mL, and the purity was 96.1%-97.2%. It only reacted with the corresponding antibody, and there was no cross-reaction between LPS of different strains.
[0034] 2. Preparation of colloidal gold and selection of optimum particle size
[0035] Use sodium citrate reduction method and tannic acid-sodium citrate reduction method to prepare 20nm, 25nm, 30nm, 40nm colloidal gold particles, observe the dispersion and uniformity results under the electron microscope, and then select the most suitable colloidal gold particle size of 20nm and 25nm to mark.
Embodiment 2
[0037] 1. Determination of the optimal labeling pH of the animal Brucella gold standard probe
[0038] Using colloidal gold gradient method and O value curve method, the optimum pH value of colloidal gold-labeled SPA was determined to be 5.2-6.8.
[0039] 2. Determination of the optimal labeling amount of the animal Brucella gold standard probe
[0040] Using the colloidal gold gradient method and the O value curve method, the optimal labeling amount of colloidal gold-labeled SPA was determined to be 20-60 μg / mL.
[0041] 3. Preparation and purification of gold-labeled probes
[0042] Take two test tubes, add 5mL 20nm and 25nm colloidal gold respectively; add 30μ1~50μl0.2mol / L K 2 CO 3 , adjust the pH value to 5.2~6.8; calculate the total amount of protein to be labeled according to the total amount of colloidal gold used for labeling to be 60μg / ml~90μg / ml, and add SPA to the colloidal gold solution drop by drop , the whole process is about 5min. Shake well for 15 minutes...
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