Preparation method of agkistrodon acutus venom detection kit based on improved double-antibody sandwich ELISA method

Abstract

The invention discloses a preparation method of an agkistrodon acutus venom detection kit based on an improved double-antibody sandwich ELISA (Enzyme-Linked Immunosorbent Assay) method, which comprises the following steps: step 1, primary purification: primarily purifying horse anti-agkistrodon acutus IgG in horse anti-agkistrodon acutus plasma by adopting a saturated ammonium sulfate method; 2, repeatedly purifying, further performing chromatographic purification on the horse anti-agkistrodon acutus IgG by adopting affinity chromatography, collecting an antibody corresponding to a target peak, and performing column chromatography purification again to obtain relatively pure horse anti-agkistrodon acutus IgG; 3, coating an antibody: taking a part of the antibody purified in the step 2 as a coating antibody, and coating the coating antibody in a 48-pore plate; step 4, labeling an antibody, taking a part of the antibody purified in the step 2 to react with horse radish peroxidase (HRP) to obtain a horse radish peroxidase labeled antibody (HRP-IgG), and taking the HRP-IgG as a sandwich antibody; and 5, packaging the kit, sealing the 48-pore plate coated in the step 3 in vacuum, and putting the 48-pore plate and the sandwich antibody in the step 4 into the kit for later use, so that the kit is prepared and can be used for detecting whether human or animal serum contains agkistrodon acutus toxin or not.

Description

technical field [0001] The invention relates to the technical field of snake venom detection, in particular to a preparation method of a Agkistrodon akistros venom detection kit based on an improved double-antibody sandwich ELISA technology. Background technique [0002] In daily life, poisonous snake bites often occur. In emergency hospitals, the type of poisonous snake is often judged based on the patient’s description of the poisonous snake or the corpse of the poisonous snake, so as to select targeted antivenom for rescue treatment. If it is impossible to determine the type of poisonous snake At that time, the previous practice was to choose multivalent antivenom for detoxification, but the potency of multivalent antivenom is low, requiring large doses of injection, and its side effects are large; now only monovalent antivenom is produced in China, fast and accurate Identifying the type of snakebite has just become a top priority. [0003] In the prior art, a Venom Dete...

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Problems solved by technology

[0002] In daily life, poisonous snake bites often occur. In emergency hospitals, the type of poisonous snake is often judged based on the patient’s description of the poisonous snake or the corpse of the poisonous snake, so as to select targeted antivenom for rescue treatment. If it is impossible to determine the type of poisonous snake At that time, the previous practice was to choose multivalent antivenom for detoxification, but the potency of multivalent antivenom is low, requiring large doses of injection, and its side effects are large; now only monovalent antivenom is produced in China, fast and accurate Identifying the type of snakebite has become a top priority

[0003] In the prior art, a Venom Detection Kit (VDK) has been used abroad for clinical use, but it is aimed at a small number of foreign snake species. Due to the difference in species, foreign test kits are not suitable for the identification of venomous snakes in China; At present, there is no rapid and accurate detection method for venomous snake bites at home and abroad, so it is very necessary to explore a simple and efficient snake venom detection method that can quickly diagnose common venomous snake bites

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