31 results about "Dot elisa" patented technology

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.

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 discloses a hybridoma cell strain secreting potato-virus-Y(PVY)-resistant monoclonal antibodies and monoclonal antibody application thereof. Pure PVY virion purified through a differential centrifugation method serves as antigens to immune BALB/c mice, cell fusion, screening and cloning are performed, one hybridoma cell strain 3B2 which is stable in passage and can secrete PVY-resistant monoclonal antibodies is obtained, and the preservation number of the hybridoma cell strain 3B2 is CGMCC No. 12001. 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 PVY coat protein of 30 kDa have a specific reaction. The 3B2 monoclonal antibodies are used for establishing ACP-ELISA, dot-ELISA and Tissue blot-ELISA detecting methods of PVY on crops, wherein the diseased leaf detecting sensitivity of the ACP-ELISA method and the diseased leaf detecting sensitivity of the dot-ELISA method reach 1:81920 and 1:10240 fold dilution (w/v, g/mL) respectively. Preparation of the PVY-resistant monoclonal antibodies and establishment of the detection methods of the PVY-resistant monoclonal antibodies provide matter and technologic support for detection and diagnosis, epidemiological analysis and scientific prevention and control of potato virus diseases.

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 provides a diagnostic kit for detecting Paragonimiasis, which contains a rabbit polyclonal antibody against the soluble antigen of Paragonia wescheri adult worm, an enzyme-labeled rabbit polyclonal antibody against the soluble antigen of Paragonimus westermani adult worm, Nitrocellulose membrane, bovine serum albumin, PBST washing solution, substrate chromogenic solution, positive control sample, negative control sample and polyethylene reaction plate. The invention also provides a preparation method of a diagnostic kit for detecting paragonimiasis. The invention adopts the double-antibody sandwich method Dot-ELISA to detect the circulating antigen of Paragonimus westermani, has high sensitivity and strong specificity, and can realize rapid diagnosis and curative effect assessment of Paragonimiasis.

Disclosed is dot-ELISA detection method of main outer-membrane protein of vibrio vulnficus. Cultured vibrio vulnficus is crashed (power 200w, beyond 5s stopping 5s) through ultrasonic, supernatant is centrifugally collected. Spotting 1.0 mu l is on a polyvinylidene fluoride film (PVDF film), after the spotting PVDF film is closed by 2% BSA (bovine serum albumin) under 37 DEG C, respectively reacted with rabbit anti vibrio vulnficus serum and goat anti-rabbit IgG whole serum marked by HRP (horseradish peroxide), finally reacted with substrate solution, macroscopic brown yellow round sedimentation is formed on the PVDF film, namely masculine result. The inventive method has detection sensitivity 15 ng/ml for main outer-membrane protein of vibrio vulnficus, is 3-5 times higher than enzyme linked immunosorbent assay (ELISA), the result is obtained after 2-3 hours, therefore vibrio vulnficus in the marine environment is detected fast and sensitively.

The invention discloses application of a hybridoma cell line for secreting a monoclonal antibody resisting rice strip mosaic virus (RSMV) and the monoclonal antibody of the hybridoma cell line. The purified RSMV is an antigen immune BALB/c mouse; a hybridoma cell line 1D4 which can stably pass and secrete the monoclonal antibody resisting the RSMV is obtained by cell fusion, screening and colonizing, and the preservation number of the hybridoma cell line 1D4 is CGMCC (China General Microbiological Culture) No.14898. The ascites indirect ELISA (Enzyme Lined Immunosorbent Assay) titer of the monoclonal antibody secreted by the hybridoma cell reaches 10<-10>, and the type and subclass of the antibody are IgG1 and kappa chains. The monoclonal antibody has specific reaction with N protein of the RSMV. An ACP-ELISA method and a dot-ELISA method for detecting the RSMV are established by taking the 1D4 as a core; the sensitivities of the two methods for detecting disease leaves respectively reach the dilution of 1 to 2621440 and the dilution of 1 to 40960 (w/v, g/mL). The obtaining of the hybridoma cell line and the monoclonal antibody thereof and establishment of a serological testing method provide material and technical support for detection and diagnosis and scientific prevention and control of virus 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 a PPV (plum pox virus)-resistant monoclonal antibody and an application of the monoclonal antibody. Prokaryotic-expression PPV coat protein istaken as an antigen to immunize a BALB/c mouse, one hybridoma cell strain 4A8 capable of secreting the PPV-resistant monoclonal antibody is obtained by cell fusion, screening and cloning, and the preservation number of the hybridoma cell strain is CGMCC No.17283. Indirect ELISA valence of monoclonal antibody ascites secreted by the cell strain reaches 10<-7>, the type and the subclass of the antibody are IgG1 and kappa light chains, and the monoclonal antibody has a specific reaction with coat protein subunit of PPV. TAS-ELISA and dot-ELISA detection methods for detecting the PPV in plum trees are established on the basis of the monoclonal antibody 4A8, wherein sensitivity of the TAS-ELISA and dot-ELISA methods for detecting sick leaves reaches 20480-fold dilution and 5120-fold dilution (w/v,g/mL). The material and technical support is provided for diagnosis and detection of PPV disease in an orchard, import and export port inspection and quarantine as well as scientific prevention and control through establishment of the preparation and detection methods of the PPV monoclonal antibody.

The invention discloses a hybridoma cell strain capable of secreting a BYDV (barley yellow dwarf virus) PAV strain monoclonal antibody and an application of the monoclonal antibody of the hybridoma cell strain. Purified BYDV PAV strain virions are used as an antigen to immunize a BALB/c mouse, one hybridoma cell strain 24G4 capable of passaging steadily and secreting the BYDV PAV strain monoclonal antibody is obtained through cell fusion, screening and cloning, and the preservation number of the hybridoma cell strain 24G4 is CGMCC No.13298. The indirect ELISA titer of monoclonal antibody ascites secreted by the hybridoma cell strain is 10<-9>, and the antibody type and subtype are IgG1 and kappa chain. ACP-ELISA and dot-ELISA methods for detecting BYDV PAV in wheat plants are established with the monoclonal antibody 24G4 as the core, and the sensitivities of the two methods for detecting infected leaves reach 1:81920 and 1:1280 fold dilution (w/v, g/mL). The hybridoma cell strain 24G4, the monoclonal antibody obtained from the same and the established serological test methods provide material and technical support for detection, diagnosis and scientific prevention and control of the viral diseases.

The invention discloses a hybridoma cell strain secreting a monoclonal antibody against rice ragged stunt viruses (RRSV) and use of the monoclonal antibody. Purified RRSV coat protein is used as an antigen to immunize a BALB/C mouse, and thus a hybridoma cell strain 4H5 which can be passed down steadily and can secrete monoclonal antibody against RRSV is obtained, wherein the collection number ofthe hybridoma cell strain 4H5 is CGMCCNo.5536. The result of Westernblot detection indicates the monoclonal antibody secreted by the hybridoma cell strain 4H5 reacts with the RRSV coat protein specifically. The indirect enzyme-linked immuno sorbent assay(ELSA) titer of the hybridoma cell strain 4H5 monoclonal antibody ascitic fluid reaches 10<-7>, the type and subtype of the antibody is IgG1 and kappa chain. According to a dot-ELISA detection method established by using the 4H5 monoclonal antibody for detecting brown rice plant hopper and RRSV in rice, when single-head brown rice plant hopperis diluted to 800mu L and affected leaves is diluted according to a weight-volume (g/mL) ratio of 1:320, viruses can still be detected. The established dot-ELISA method can detect brown rice plant hopper and RRSV in rice accurately, specifically and sensitively. The preparation of the monoclonal antibody against RRSV and the establishment of the detection method thereof provide technical and material supports for diagnosing, predicting and forecasting and scientifically preventing and controlling rice virus diseases.