Immunoaffinity column for detecting vomitoxin and application of immunoaffinity column
A technology of vomitoxin and immunoprophylaxis, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of poor experimental sensitivity and repeatability, serious injury to operators, cumbersome operation process, etc., and achieve short pretreatment time, simple storage, The effect of high sensitivity
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Embodiment 1
[0035] Example 1 Preparation of immunoaffinity column for detection of vomitoxin
[0036] The preparation method of this immunoaffinity column mainly comprises the following steps:
[0037] 1) Prepare the solid phase medium coupled with vomitoxin antibody, i.e. CNBr-Sepharose-4B-Ab
[0038] 2) Load the solid phase medium into the column, and assemble the immunoaffinity column according to the combination and connection method of the affinity column.
[0039] The following step-by-step detailed description:
[0040] 1. Preparation of vomitoxin hapten
[0041] Mix 0.10g vomitoxin in 2mL dimethyl sulfoxide (DMSO), slowly add dropwise 0.1mL 1,3-propanediamine and 0.1mL pyridine in 2mL DMSO mixture at 60°C, continue the reaction after the dropwise addition is complete After 12h, the solvent and unreacted propylenediamine were removed by rotary evaporation, and the propylenediamine monocondensate of vomitoxin was quantitatively obtained. The synthetic route is as follows: figur...
Embodiment 2
[0059] Example 2 Detection of vomitoxin residues in samples
[0060] 1. Sample pretreatment
[0061] Homogenize the sample with a homogenizer; weigh 2.0±0.05g of the sample into a 50ml polystyrene centrifuge tube, add 16ml of deionized water, vortex with a vortexer for 5min, or shake on a shaker for 20min, above 3000g, room temperature (20 -25℃-77 / 68℉) centrifuge for 5min; filter all supernatant with qualitative filter paper for later use;
[0062] 2. Operation steps of sample passing through the column
[0063] Connect the immunoaffinity column under a 10ml syringe, and connect the affinity column product with an adapter; accurately draw 8ml (equivalent to 1g sample) of the test solution into the syringe, and use the syringe to control the flow rate of the test solution for 2 seconds Each drop passes through the affinity column; rinse the affinity column once with 5ml of deionized water, use a syringe to control the flow rate to 0.5-1 second per drop, and drain the liquid i...
Embodiment 3
[0064] Embodiment 3 sample detection example
[0065] 1. Blank determination of negative samples: the general matrix category includes (cereals, finished feed).
[0066] Take the blank samples of the above-mentioned various matrices (corn, soybean meal, wheat bran, distiller's grains, batch materials, concentrated materials), purify them through the vomitoxin immunoaffinity column, and perform instrumental detection to obtain the following results (see Table 1), each Samples were measured in triplicate.
[0067] Table 1 Various blank sample detection results
[0068]
[0069] The results show that the blank value detected by various matrices is far less than the detection limit of 1 μg / kg required by the state, indicating that the affinity column can be applied to the detection of samples of various matrices, and different sample matrices will not cause blank interference to the detection.
[0070] 2. Determination of accuracy and precision: Add vomitoxin at a concentrati...
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