Indirect ELISA (Enzyme-Linked Immunosorbent Assay) method and kit for detecting haemophilus parasuis antibodies

A technology of Haemophilus suis and antibodies, applied in biological testing, botany equipment and methods, biochemical equipment and methods, etc., can solve the problems of multiple components, complex influencing factors, difficult application, etc., and achieve high biological safety and wide Application prospects, reproducible and fast effects

Inactive Publication Date: 2011-02-09
广东省农业科学院兽医研究所
View PDF1 Cites 22 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the PCR and Southern hybridization methods based on the detection of its DNA are difficult to apply in practice, and CF also has the disadvantages of many components involved in the reaction, complex influencing factors, and cumbersome operation steps. IHA and ELISA methods are currently the main HPS methods explored by scholars at home and abroad. Antibody detection method, the difference is the preparation method of the antigen, and Feng Xiaoming (2009), Shi Bi

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Indirect ELISA (Enzyme-Linked Immunosorbent Assay) method and kit for detecting haemophilus parasuis antibodies
  • Indirect ELISA (Enzyme-Linked Immunosorbent Assay) method and kit for detecting haemophilus parasuis antibodies
  • Indirect ELISA (Enzyme-Linked Immunosorbent Assay) method and kit for detecting haemophilus parasuis antibodies

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0030] Example 1

[0031] 1. Cloning, expression and purification of OMP P5 protein gene of Haemophilus parasuis

[0032] 1. PCR primer design and synthesis

[0033] (1) According to the gene sequence of HPS OMP P5 (gene accession number is CP001321), a pair of specific primers was designed with the biological software Premier Primer 5.0 to amplify the whole gene of HPS OMP P5. Primers were synthesized by Shanghai Sangon Bioengineering Technology Service Co., Ltd. Sequence of upstream primer HPS P1: 5′-GCC GATATC GCTCCACAAGCTAACACT-3' (the underlined part is the EcoRV restriction site); the sequence of the downstream primer HPS P2: 5'-CGC GGATCC TTACATAGAAACTTCTTTTGAACCT-3' (the underlined part is the BamHI restriction site).

[0034] 2. Cloning and expression vector construction of HPS OMP P5 gene

[0035] Using the HPS bacterial liquid of clinical isolates as a template, the designed specific primers P1 / P2 were used for amplification (pre-denaturation at 95 °C for 5 ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses indirect ELISA (Enzyme-Linked Immunosorbent Assay) method and kit for detecting haemophilus parasuis antibodies. The indirect ELISA method comprises the steps of: taking a haemophilus parasuis OMPP5 protein as an envelope antigen; wherein the judgment standard is as followed: OD450 value of to-be-detected serum is greater than 0.375, OD450 value of the to-be-detected serum/OD450 value of standard negative serum is greater than or equal to 2.1. The kit for realizing the method comprises an envelope buffer solution, confining liquid, a washing buffer solution, an antibody diluent, a chromogenic substrate solution and a stop solution, wherein the envelop antigen used in the method is the haemophilus parasuis OMPP5 protein. Proved by a specificity test, a blocking-up experiment, a repeatability test and clinic application detection, the invention has the characteristics of excellent specificity, high sensitiveness, good repeatability, rapidness, simpleness and accuracy, and can be used in clinic large-scale detection of haemophilus parasuis antibodies and epidemiological survey.

Description

technical field [0001] The invention relates to the technical field of molecular biology, animal bacteriology and animal infectious disease detection, in particular to an indirect ELISA method and a kit for detecting Haemophilus parasuis antibodies. Background technique [0002] Haemophilus parasuis disease is one of the important causes of morbidity and mortality in weaned piglets. Haemophilus parasuis (HPS), as a common bacterium in the upper respiratory tract of pigs, invades the body under certain conditions and can cause severe systemic diseases (Glasser's disease), such as fibrinous polyserositis, joint inflammation and meningitis are the main features. This disease has been considered to be a sporadic, stress-induced disease in piglets in the past. However, in recent years, with the development of pig farming industry, more and more high-health pig herds or SPF pig herds appear. If these pigs are in an unimmunized state, once HPS invades, they will be seriously infe...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12N15/31G01N33/68C07K14/195C12N15/70
Inventor 李春玲王贵平陈善真宋帅贾爱卿
Owner 广东省农业科学院兽医研究所
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products