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Enzyme-linked immunosorbent kit for inspecting porcine immunoglobulin G and application thereof

An enzyme-linked immunosorbent reagent and immunoglobulin technology, which is applied in measurement devices, color/spectral property measurement, instruments, etc., can solve the problems of cumbersome operation, difficult systematization and standardization of technology, and complicated equipment and equipment.

Inactive Publication Date: 2009-11-04
贵州谱尼测试技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the difficulty of systematization and standardization of the technology used in the above method, complex equipment and cumbersome operation, etc., it is not suitable for on-site inspection and screening of a large number of samples.

Method used

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  • Enzyme-linked immunosorbent kit for inspecting porcine immunoglobulin G and application thereof
  • Enzyme-linked immunosorbent kit for inspecting porcine immunoglobulin G and application thereof
  • Enzyme-linked immunosorbent kit for inspecting porcine immunoglobulin G and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] The preparation of embodiment 1 kit components

[0057] 1. Preparation of Monoclonal Antibody

[0058] a. Animal immunity

[0059] The immunogen was injected into Balb / c mice with an immunization dose of 100 μg / mouse to produce polyclonal antibody serum.

[0060] b. Cell fusion and cloning

[0061] After the measurement result of mouse serum was higher, the splenocytes were taken and fused with SP2 / 0 myeloma cells at a ratio of 9:1, and the cell supernatant was measured by indirect competitive ELISA, and the positive wells were screened. Positive wells were cloned by limiting dilution until hybridoma cell lines secreting monoclonal antibodies were obtained.

[0062] The porcine immunoglobulin G monoclonal hybridoma cell line C-4-2CGMCC No.3110 was obtained through screening. The monoclonal hybridoma cell line of porcine immunoglobulin G can produce unlimited amount of porcine immunoglobulin G specific antibody, and the specificity of the antibody is for porcine immu...

Embodiment 2

[0076] Embodiment 2 detects the formation of the enzyme-linked immunosorbent assay kit of porcine immunoglobulin G

[0077] The ELISA kit for detecting porcine immunoglobulin G is set up to include the following components:

[0078] (1) A microtiter plate coated with porcine immunoglobulin G-coupled antigen;

[0079] (2) Goat anti-mouse anti-antibody labeled with horseradish peroxidase;

[0080] (3) Porcine immunoglobulin G monoclonal antibody working solution;

[0081] (4) 6 bottles of porcine immunoglobulin G standard solution, the concentrations are 0mg / L, 0.2mg / L, 0.6mg / L, 1.8mg / L, 5.4mg / L, 16.2mg / L;

[0082] (5) The substrate chromogenic solution is composed of A liquid and B liquid, the substrate chromogenic liquid A liquid is carbamide peroxide, and the substrate chromogenic liquid B liquid tetramethylbenzidine;

[0083] (6) The stop solution is 2mol / L hydrochloric acid;

[0084] (7) The concentrated washing solution is 0.5%-1.0% Tween 20 and 0.1-0.3‰ thimerosal pre...

Embodiment 3

[0086] The detection of porcine immunoglobulin G in the sample of embodiment 3

[0087] 1. Sample pretreatment

[0088] Weigh 20 mg of the sample to be tested and dissolve it in 40 mL of deionized water to make the final solution concentration 0.5 mg / mL. After dissolving, shake and mix well, and then fully mix it with the diluted sample complex solution at a ratio of 1:49 spare. Take 50 μl for analysis.

[0089] 2. Detection with kit

[0090] Add 50 μl of porcine immunoglobulin G standard solution or sample solution to the microwell of the microplate coated with porcine immunoglobulin G antigen, then add 50 μl of porcine immunoglobulin G monoclonal antibody working solution, and then add horseradish Oxidase-labeled goat anti-mouse anti-antibody working solution 50μl, seal the plate with a cover plate, react in a 37°C incubator for 30min, pour out the liquid in the well, add 250μl washing solution to each well, pour out the liquid in the well after 30s, and so on Repeat the...

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Abstract

The invention provides an enzyme-linked immunosorbent kit for inspecting porcine immunoglobulin G, comprising an ELISA plate which is coated with coating antigen, an enzyme label, porcine immunoglobulin G specific antibody working liquid (being contained when the antigen is coated on the ELISA plate and the enzyme label is enzyme labeling antibody or antibody is coated on the ELISA plate and the enzyme label is enzyme labeling antigen), porcine immunoglobulin G standard product solution, substrate color development solution, stop solution, concentrated washing liquid and concentrated complex solution. The invention further discloses a method which applies the enzyme-linked immunosorbent kit for inspecting the porcine immunoglobulin G, and the method comprises the steps of firstly carrying out the pre-treatment on a sample, then using the kit for inspecting and finally analyzing the inspection result. The provided enzyme-linked immunosorbent kit can be used for inspecting the content ofthe porcine immunoglobulin G in porcine plasma protein powder and other samples, the operation is simple, the cost is low, and the enzyme-linked immunosorbent kit can be monitored on-site and is appl icable in screening mass samples.

Description

technical field [0001] The invention relates to an ELISA detection technology, in particular to an ELISA reagent kit for detecting porcine immunoglobulin G, which is particularly suitable for the determination of porcine immunoglobulin G in porcine plasma protein powder. technical background [0002] As a new type of protein feed, plasma protein powder has the characteristics of comprehensive nutrition, easy digestion and absorption, good palatability, rich in immunoglobulin, etc., and is widely added in piglet feed. According to the sources and processing methods of blood raw materials, there are mainly the following categories: porcine plasma protein powder (spray-dried plasma protein, SDPP), low ash porcine plasma protein powder (LAPP), sow plasma protein powder (SDSPP) and bovine plasma protein powder (SDBP), the effect is roughly the same, among which SDPP is the most commonly used, so in general, plasma protein powder refers to SDPP. [0003] A large number of studies...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/577G01N33/535G01N21/31
Inventor 万宇平高东微王雅静马孝斌刘中勇蔡克周余厚美陈永红李明
Owner 贵州谱尼测试技术有限公司
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