Detecting method for quick detecting veterinary medicinal residue in food and agriculture and sideline products
A technology for agricultural and sideline products and veterinary drug residues, which is applied in the preparation of test samples, measuring devices, instruments, etc., can solve the problems of narrow application range, poor specificity, and complicated operation.
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Embodiment 1
[0034]Pretreatment of honey samples: Take 1g of honey, put it into a centrifuge tube, add 2mL of distilled water to dissolve; add 2mL of ethyl acetate to shake for 10min; centrifuge at 3000g at room temperature for 10min; In a test tube, blow dry with N2 at 60°C; dissolve the residue with 0.5mL buffer 1; take 50uL for analysis. Put the processed sample into a centrifuge tube, add 6 mL of ethyl acetate and shake for 10 min; centrifuge at 3000 g at room temperature for 10 min; take out 2 mL of the upper layer of ethyl acetate (equivalent to 1 g of sample) and dry it under N2 at 60 °C.
[0035] Preparation of washing liquid: Tris54g, boric acid 27.5g, dissolve with a small amount of DD-H2O, add 20ml of 0.5mol / L EDTA-Na2 (PH8.0), add DD-H2O to dilute to 1000ml. Add 200ml of saturated (NH4)2SO4 solution, 40ml of Tween-20 and 480ml of DD-H2O, and mix well.
[0036] Preparation of reagent biotinylated antibody: take 1g of biotin, suspend it in 12ml N,N dimethylformamide DMF, add 0.6...
Embodiment 2
[0043] The pretreatment of sample and the preparation of washing solution are the same as in Example 1
[0044] Preparation of reagent biotinylated antibody: Take 1g of biotin, suspend it in 12ml N,N dimethylformamide DMF, add 0.6g N-hydroxysuccinimide HOSU and 0.8g dicyclohexylcarbodiimide DCC was placed in an airtight container and magnetically stirred overnight at room temperature. The filtrate was spun to dryness. Add 10ml of diethyl ether for washing, and then add 200ml of isopropanol for crystallization to obtain white powdery activated biotin crystals. Use DMF to make BNHS into a 1mg / mL solution; dilute the antibody to be labeled with 0.10mol / L sodium bicarbonate solution with a pH value of 9.0 to 2mg / ml, and after dialysis, use 1mg / ml BNHS: antibody to be labeled = 1 : 10 and mix with occasional shaking for 3 hours. Add 0.05mol / L, pH7.4 PBS solution into the dialysis bag, dialyze overnight at 4°C, and dilute to the working concentration when used.
[0045] Reagent ...
Embodiment 3
[0049] The pretreatment of sample and the preparation of washing solution are the same as in Example 1
[0050] Preparation of reagent biotinylated antibody: take 1g of biotin, suspend it in 12ml N,N dimethylformamide DMF, add 0.6g N-hydroxysuccinimide HOSU and 0.8g dicyclohexylcarbodiimide DCC , placed in an airtight container, and magnetically stirred overnight at room temperature. The filtrate was spun to dryness. Add 10ml of diethyl ether for washing, and then add 200ml of isopropanol for crystallization to obtain white powdery activated biotin crystals. Use DMF to make BNHS into a 1.2 mg / mL solution; dilute the antibody to be labeled with 0.12 mol / L sodium bicarbonate solution with a pH value of 9.2 to 32 mg / ml, and after dialysis, use 1 mg / ml BNHS: Antibody to be labeled = Mix at a ratio of 1:15 with occasional shaking for 5 hours. Add 0.1mol / L, pH7.6 PBS solution into the dialysis bag, dialyze overnight at 4°C, and dilute to the working concentration when used.
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