31 results about "Dot elisa" patented technology

A monoclonal antibody-based Dot-ELISA assay for the rapid detection of animal viruses such as avian influenza virus. The assay includes applying a specimen suspected of containing an animal virus on a porous membrane and treating the specimen with a solution of citric acid or lactic acid and a solution containing a mucolytic agent and a detergent. The treated specimen is then contacted with a primary monoclonal antibody for detecting the virus. If present, the primary moncolonal antibody bind with an antigen of the animal virus specimen. The specimen is contacted with an anti-monoclonal antibody conjugate (secondary antibody) and incubated to facilitate binding of the antigen-bound monoclonal antibody to the conjugate. The bound conjugate and antigen-bound monoclonal antibody is contacted with a coloring reagent to allow visual detection of the presence of the animal virus in the specimen.

The invention relates to the field of biotechnology, and specifically discloses a canine single-chain antibody and its construction method and application. The invention successfully amplifies the canine heavy chain variable region VH and light chain variable region VL genes by designing degenerate primers for the variable region of the canine antibody. On this basis, the canine single-chain antibody scFv library was successfully constructed using phage display technology. The constructed scFv single-chain antibody library has diversity and large enough library capacity. The present invention uses the dog DEA1.1 blood group antigen to screen the scFv single-chain antibody library to obtain 3 single-chain antibodies capable of binding to the dog DEA1.1 blood group antigen by Dot-ELISA method. Among them, Clone16 has strong binding properties. The pET‑32a‑Clone16 recombinant protein was constructed based on the Clone16 gene for prokaryotic expression, and the purified antibody had binding activity. It provides a solid foundation for the preparation of canine DEA1.1 blood group identification reagent.

The invention discloses a hybridoma cell strain capable of secreting an anti-Southern-bean-mosaic-virus monoclonal antibody and an application of the monoclonal antibody thereof. Purified Southern bean mosaic virus (SBMV) particles serve as antigen immunization BALB / c mice, and are subjected to cell fusion, cell screening and cell cloning, the hybridoma cell strain 19H9 capable of conducting stable passage and secreting the anti-SBMV monoclonal antibody is obtained, and the collection number is CGMCC No.12004. The ascites indirect ELISA titer of the monoclonal antibody secreted by the hybridoma cell strain is 10<-7>, and the type and the subgenera of the antibody are IgG1 light chains and kappa light chains. The monoclonal antibody and SBMV are subjected to a specific reaction. An ACP-ELISA method for detecting the SBMV in leguminous plants and a dot-ELISA method for detecting the SBMV in leguminous plants are built with the monoclonal antibody 19H9, and the diseased leaf detection sensitivity of the ACP-ELISA method and the dot-ELISA method is 1:81,920 w / v and 1:5,120 g / ml. The hybridoma cell strain 19H9, the obtained monoclonal antibody and the built serological detecting method provide substance supports and technical supports for diagnosing, detecting and scientific preventing and controlling of the virus disease.

The invention discloses a dot enzyme-linked immunosorbent assay (ELISA) method for detecting bemisia tabaci carrying a tomato yellow leaf curl virus and application of the method. Field bemisia tabaci is collected, pretreated and grinded, monoclonal antibody of the tomato yellow leaf curl virus is used, the dot-ELISA method which is used for detecting virus transmission medium bemisia tabaci is established, a quick kit is developed and researched, and the tomato yellow leaf curl virus which is carried by the bemisia tabaci is detected. By the aid of the method, the virus carrying rate of the field bemisia tabaci can be surveyed in a large scale, and the field tomato yellow leaf curl disease outbreak trend is predicted. The method is quick, efficient, sensitive, special, high in flux and simple to operate. The tomato yellow leaf curl disease can be diagnosed quickly, predicted and controlled scientifically.

The invention provides an aeromonas hydrophila recombinant aerolysin Dot-enzyme-linked immuno sorbent assay (ELISA) detection method and belongs to the technical field of biology. A nitrocellulose (NC) film is taken as a solid-phase carrier, aeromonas hydrophila recombinant aerolysin rabbit antiserum is taken as a primary antibody, and a hypothalamic regulatory peptide (HRP) goat-anti-rabbit IgG is taken as a secondary antibody. The method can measure a plurality of samples, can obviously shorten the time for detecting the samples, can be operated easily and rapidly, can meet the requirement under a non-laboratory condition simultaneously, and can be rapidly and conveniently used for detecting pathogenic aeromonas hydrophila.

The invention discloses a hybridoma cell strain secreting citrus yellow vein clearing viru(CYVCV)-resistant monoclonal antibodies and monoclonal antibody application thereof. Coat protein of prokaryotic-expression CYVCV serves as antigens to immune BALB / c mice, cell fusion, screening and cloning are performed, one hybridoma cell strain 18H5 which is stable in passage and can secrete CYVCV-resistant monoclonal antibodies is obtained, and the preservation number of the hybridoma cell strain 18H5 is CGMCC No. 12003. The ascites indirect ELISA titer of the monoclonal antibodies secreted by the cell strain reaches 10<-7>, according to the antibody type and the subclass, the monoclonal antibodies are composed of IgG1 and kappa light chains, and the monoclonal antibodies and CYVCV coat protein subunits of 32 kD have a specific reaction. The 18H5 monoclonal antibodies are used for establishing TAS-ELISA, dot-ELISA and Tissue blot-ELISA detecting methods of CYVCV in citrus trees, wherein the diseased leaf detecting sensitivity of the TAS-ELISA method and the diseased leaf detecting sensitivity of the dot-ELISA method reach 1:2560 and 1:20480 fold dilution (w / v, g / mL) respectively. Preparation of the CYVCV-resistant monoclonal antibodies and establishment of the detection methods of the CYVCV-resistant monoclonal antibodies provide matter and technologic support for detection and diagnosis, epidemiological analysis and scientific prevention and control of citrus virus diseases.

The invention discloses a hybridoma cell strain capable of secreting a wheat dwarf virus resisting monoclonal antibody and a monoclonal antibody application of the strain. WDV (wheat dwarf virus) CP recombinant protein in prokaryotic expression is used as an antigen immune for BALB / c mice, the hybridoma cell strain 23F4 capable of stably going down to the future generation and secreting the WDV antibody is obtained after cell fusion, screening and cloning, and the collection number of the strain is CGMCC No. 13297. The abdominal dropsy indirect ELISA (enzyme-linked immuno sorbent assay) valence of the antibody secreted by the hybridoma cell strain reaches 10<-9>, and the antibody type and subclass are IgG1 and kappa chains. The antibody and WDV coat protein have specific reaction. The 23F4 antibody is used as a core for building ACP-ELISA and dot-ELISA methods for detecting WDV in a wheat plant, and diseased leaf detection sensitivity of the two methods reach 1:163840 and 1:5120 times of dilution (w / v, g / mL) respectively. Acquisition of the hybridoma cell strain and the antibody thereof and building of serological detection methods provide substantial and technical support for virus disease detection, diagnosis and scientific prevention and control.

The invention discloses a use of a hybridoma cell line secreting a monoclonal antibody against rice gall dwarf viruses and a use thereof. A BALB / c mouse is immunized by a prokaryotically expressed rice gall dwarf virus (RGDV) CP recombinant protein as an antigen, and a hybridoma cell line 11B6 capable of stable passage and secretion of an anti-RGDV monoclonal antibody is obtained by cell fusion, screening and cloning and has a preservation number of CGMCC No. 14897. The indirect ELISA titer of the ascites monoclonal antibody secreted by the hybridoma cell line is 10<-8>. The antibody type andsubclass are IgG1 and kappa chains. The monoclonal antibody can react specifically with the RGDV coat protein. The ACP-ELISA and dot-ELISA methods for detecting RGDV are established through a 11B6 monoclonal antibody as the core. The sensitivity of the two methods for detecting diseased leaves is 1:327 680-fold dilution (w / v, g / mL) and 1:20 480-fold dilution (w / v, g / mL). The acquisition of the hybridoma cell line and its monoclonal antibody and the establishment of serological detection methods provide material and technical support for the detection, diagnosis and scientific prevention and control of the viral disease.

The present invention provides a recombinant prokaryotic expression protein antigen for detecting autochthonous trichinella spiralis antibody and its preparation method. Said product can be used as detection antigen of indirect-ELISA and Dot-ELISA method for detecting autochthonous trichinella spiralis antibody, also can be used as gene engineering subunit vaccine for preventing autochthonous trichinella spiralis infection. The preparation process of said product includes the following steps: (1) designing specific primer for amplifying TNPG gene; (2), utilizing PCR amplification to obtain TNPG gene segment; (3), cloning TNPG gene and sequencing, and making identification; (4), directionally subcloning TNPG gene to pET-3a prokaryotic expression vector; (5), induced expression of TNPG gene in colibacillus; and (6), affinity chromatography purification of TNPG gene expression protein.

The invention discloses a hybridoma cell strain secreting monoclonal antibody against watermelon mosaic virus and application of the monoclonal antibody. Purified virus particles of watermelon mosaic virus (WMV) are used as an antigen to immunize BALB / c mice; through cell fusion, screening and cloning, one strain of hybridoma cell strain 1C5 capable of stable passage and secreting anti-WMV monoclonal antibodies is obtained; and the strain has a preservation number CGMCC No.9340. The monoclonal antibody secreted from the hybridoma cell strain has ascites indirect ELISA titer of 10 <-7>, and the antibody type and subtype are IgG1, kappa chain. The monoclonal antibody has specificity reaction with CMV. The 1C5 antibody is used to establish a dot-ELISA virus method for detection of WMV in aphid and in white pumpkin. The hybridoma cell strain 1C5, monoclonal antibody thereof and the establishment of the related serological detection method provide technical and material support for the diagnosis, prediction and scientific prevention and control of the virus.

The invention discloses an indirect ELISA (enzyme-linked immunosorbent assay) based detection kit for SrMV (sorghum mosaic virus) in sugarcane. The indirect ELISA based detection kit mainly comprises a rabbit-anti polyclonal antibody specifically recognizing SrMV as well as an HRP (horseradish peroxidase)-labeled goat anti-rabbit IgG standard, wherein the polyclonal antibody is prepared from polypeptide in an N-end conserved region of P3 protein of SrMV of the sugarcane as an antigen. Accordingly, a corresponding indirect ELISA based detection method is established and used for detecting SrMV in the sugarcane. Compared with the prior art, the Dot-ELISA based detection kit and method have the advantages of high sensitivity, good specificity, low cost, high throughput, rapid detection and the like, is both suitable for rapid detection of a small quantity of samples in field and high-throughput rapid detection of a large quantity of samples in a laboratory.

The invention discloses a hybridoma cell strain excreting a monoclonal antibody (MAb) resisting a rice blackstreaked dwarf virus (RBSDV) and the application of the MAb. A coat protein (CP) of the RBSDV is expressed into antigen immune BALB / c mouse by a prokaryotic expression method, and the hybridoma cell strain 5G1 which can stably passage and excrete the MAb resisting the RBSDV is obtained through cell fusion, selection and cloning, wherein the preserving number is CGMCC No. 5537. Ascitic indirect enzyme-linked immuno sorbent assay (ELISA) valence of the 5G1 MAb is over 10<-6>, and the antibody type and the subtype are IgG1 and kappa chain. The antibody excreted by the cell strain and the coat protein of the RBSDV have specific immunobinding reaction. By the dot-ELISA detection method for detecting rice planthopper and the RBSDV in rice and established by using the 5G1 MAb as a core, the virus can be still detected when a single-head small brown rice planthopper is diluted to 1,600 microlitres and sick leaves are diluted (w / v, g / mL) in the proportion of 1:160. The preparation of the MAb resisting the RBSDV and the establishment of the detection method thereof provide technical and material support for the diagnosis, forecast and scientific prevention and control of rice viruses.