A double-antibody sandwich enzyme-linked immunosorbent assay for the detection of Bacteroides oryzae
A technology of streak streak and bacteria, which is applied in the field of immunoassay, can solve the problem that there is no effective control method for bacterial streak bacterium of rice, etc., and achieves the effects of simple pretreatment process, good stability and good specificity.
Active Publication Date: 2016-06-22
JIANGNAN UNIV
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Problems solved by technology
[0004] Since there is no effective control method for rice bacterial stripe disease in agricultural production at present, and the seed carrier is an important source of initial infection and transmission route of the disease, therefore, early detection of the pathogenic bacteria on the seeds is important for preventing the disease. It is of great significance to take corresponding prevention and control measures to guide production and reduce economic losses
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[0025] The present invention is further illustrated by the following examples.
[0026] Instruments: TGL-40B desktop low-speed centrifuge, Shanghai Anting Scientific Instrument Factory; KFLOW pure water machine, Kevron Company; ZD-9556 horizontal shaker, Taicang Science and Education Equipment Factory; 96-well 8×12 detachable microplate, Xiamen Yijiamei Laboratory Equipment Co., Ltd.; MuLtiskaMks microplate reader, ThermoLabsystems; adjustable pipette, ThermoLabsystems; vortex mixer, Shanghai Huxi Instrument Analysis Factory.
[0027] Reagents: Tetramethylbenzidine (TMB), Shanghai Jingchun Industrial Co., Ltd.; other reagents are analytical reagents.
[0028] Specific steps are as follows:
[0029] (1) Monoclonal antibody preparation:
[0030] a. Experimental animals: 5 BALB / c mice aged 8 weeks were selected for immunization;
[0031] b. Antigen configuration: Dilute the immunogen with normal saline to make 1×10 8 cfu / mL solution;
[0032] c. Emulsification: emulsify the ...
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Abstract
The invention relates to a double-antibody sandwich enzyme-linked immunosorbent method for detecting rice bacterial streak bacterium, which belongs to the technical field of immune analysis. In the present invention, mice are immunized with the rice bacterial spot pathogen to obtain 7 monoclonal antibodies against the rice bacterial stripe pathogen. The horseradish peroxidase was marked separately and paired with rice bacterial spot bacterial cells in pairs. 3D7 (monoclonal cell line No. J) was used as the coating antibody, 4H6 (monoclonal cell line No. K) was used as the enzyme-labeled antibody, and rice bacterial spot bacterial cell was used as the standard to establish the sandwich of rice bacterial spot disease ELISA method. The present invention adopts the monoclonal antibody with highly uniform physical and chemical properties, good specificity, and can be prepared in large quantities. The sandwich method established has high sensitivity and low cost. Fusarium blight, Bacterial wilt of corn, and Pseudomonas syringae pv. syringae have no cross-reactivity, providing a fast and efficient analytical method for the detection of bacterial streak disease in rice seeds.
Description
technical field [0001] The invention relates to a double-antibody sandwich enzyme-linked immunosorbent method for detecting rice bacterial streak bacterium, belonging to the technical field of immune analysis. Background technique [0002] Bacterial spot disease of rice, scientific name: Xanthomonasoryzaepv.oryzicola (Xooc) ,English name bacterial leaf streak of rice , belonging to the order Xanthomonas, the family Xanthomonas, and the genus Xanthomonas. Rice bacterial streak disease was first discovered in the Philippines in 1918, and now it occurs in Southeast Asian countries and central Africa. It mainly occurs in southern rice-producing areas in my country. Rice bacterial streak disease mainly damages rice leaves, and typical streak symptoms can appear at the seedling stage, and its occurrence has the characteristics of epidemic, explosive and destructive. Bacterial spot disease of rice is transmitted over long distances by infecting seeds and transporting seeds. Th...
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IPC IPC(8): G01N33/577G01N33/531
CPCG01N33/532G01N33/56911G01N2333/195
Inventor 匡华孔德昭胥传来徐丽广马伟刘丽强宋珊珊吴晓玲
Owner JIANGNAN UNIV