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Protocol of blocking ELISA for differentiating antibodies of swine transmissible gastroenteritis virus and porcine respiratory coronavirus
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SN/T 1697-2006
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Basic data | Standard ID | SN/T 1697-2006 (SN/T1697-2006) | | Description (Translated English) | Protocol of blocking ELISA for differentiating antibodies of swine transmissible gastroenteritis virus and porcine respiratory coronavirus | | Sector / Industry | Commodity Inspection Standard (Recommended) | | Classification of Chinese Standard | B41 | | Classification of International Standard | 11.220 | | Word Count Estimation | 6,675 | | Date of Issue | 1/26/2006 | | Date of Implementation | 2006-08-16 | | Quoted Standard | GB/T 6682 | | Regulation (derived from) | ?Industry Standard Announcement 2006 No.4 (Total No.76); National-Quality-Inspection-accreditation (2010) 290; Industry Standard Filing Announcement 2010 No.10 (Total No.130) | | Issuing agency(ies) | General Administration of Customs | | Summary | This standard specifies the porcine transmissible gastroenteritis (TGE) virus blocking ELISA antibody identification test operations technical requirements. This standard applies to porcine transmissible gastroenteritis virus, porcine respiratory coronavirus identified. | SN/T 1697-2006: Protocol of blocking ELISA for differentiating antibodies of swine transmissible gastroenteritis virus and porcine respiratory coronavirus
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Protocol of blocking ELISA for differentiating antibodies of swine transmissible gastroenteritis virus and porcine respiratory coronavirus
People's Republic of China Entry-Exit Inspection and Quarantine Industry Standard
Porcine transmissible gastroenteritis virus and porcine respiratory coronavirus
Antibody Blocking ELISA Identification Test Procedure
Released on.2006-01-26
Implementation of.2006-08-16
People's Republic
The General Administration of Quality Supervision, Inspection and Quarantine issued
Foreword
Appendix A, Appendix B and Appendix C of this standard are normative appendices.
This standard is proposed and managed by the National Certification and Accreditation Administration.
This standard was drafted. Guangdong Entry-Exit Inspection and Quarantine Bureau of the People's Republic of China.
The main drafters of this standard. Luo Changbao, Chen Ru, Liu Linlin.
This standard is the first industry standard for entry-exit inspection and quarantine.
Porcine transmissible gastroenteritis virus and porcine respiratory coronavirus
Antibody Blocking ELISA Identification Test Procedure
1 Scope
This standard specifies the ELISA test for porcine transmissible gastroenteritis (TGE) virus and porcine respiratory coronary (PRC) virus antibody blocking ELISA.
Operational technical requirements.
This standard applies to the identification of porcine transmissible gastroenteritis virus and porcine respiratory coronavirus.
2 Normative references
The terms in the following documents become the terms of this standard by reference to this standard. All dated references, followed by all
Modifications (not including errata content) or revisions do not apply to this standard, however, parties to agreements based on this standard are encouraged to study
Is it possible to use the latest version of these files? For undated references, the latest edition applies to this standard.
GB/T 6682 Analytical laboratory water specifications and experimental methods
3 materials
3.1 Viruses and cells
ST cells were grown in 96-well cell culture plates to form a monolayer and infected with the TGE virus Miller6 strain. Cytopathic rate after 24h
Fix cells at no more than 10%.
3.2 Fixed cells
The cell plate was washed once with PBS (pH 7.4) for 1 to 2 times, then fixed with 80% acetone (precooled -20 ° C) for 15 min, and the acetone was discarded.
Dry in a wind cabinet for 10 min, then store at -20 °C for later use.
3.3 antibody
Biotinylated anti-TGE virus, A site monoclonal antibody, anti-TGE virus D site monoclonal antibody.
4 method of operation
4.1 Remove the plate from -20 °C and set at room temperature. After blocking the non-specific site for 2 h at room temperature (about 25 °C) with PBST (see Appendix A), remove it.
PBST was patted dry on a paper towel and 150 μL of PBST was added to all virus-infected wells.
4.2 Add 50 μL of positive serum to A1 and A2, negative serum B1 and B2 (ie, double well), and other tested serums are listed in Appendix B.
4.3 Dilute the sample cell plate at 4 ° C overnight.
4.4 Drain the diluted sample solution and do not rinse.
4.5 Add 100 μL of working concentration biotinylated TGEA site monoclonal antibody to each well.
4.6 For identification purposes, each sample is diluted in two plates, each plate should be operated separately, using different monoclonal antibodies, separate
Count, and contain their respective negative and positive serum.
4.7 At room temperature for 2 h or 4 ° C overnight.
4.8 Wash three times with PBST.
4.9 Add 100 μL of working concentration horseradish peroxidase-labeled avidin to each well and incubate for 30 min to 40 min at room temperature.
4.10 Wash PBST three times.
4.11 Add 100μLABTS to each well (see Appendix C).
4.12 After 30 min at room temperature, 100 μL of sodium 5% sodium dodecyl sulfate (mass concentration) was added to stop the reaction.
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