GB/T 14926.28-2026 Laboratory animal—Method for examination of minute virus of mice English, Anglais, Englisch, Inglés, えいご
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ICS 13.220.10
CCS H 57
National Standard of the People's Republic of China
GB/T 14926.28-2026
Replaces GB/T 14926.28-2001
Laboratory animal - Method for examination of minute virus of mice
实验动物 小鼠细小病毒检测方法
Issue date: 2026-01-28 Implementation date: 2027-02-01
Issued by the General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China
the Standardization Administration of the People's Republic of China
Contents
Foreword
Introduction
1 Scope
2 Normative References
3 Terms and Definitions
4 Principle
5 Reagents and Equipment
6 Detection Methods
7 Result Interpretation
8 Result Reporting
Laboratory Animals — Detection Method for Minute Virus of Mice
1 Scope
This document describes detection methods for Minute Virus of Mice (MVM).
This document applies to the detection of MVM in laboratory mice, or in experimental inocula and environmental samples from laboratory animal facilities.
2 Normative References
The following documents are referred to in the text in such a way that some or all of their content constitutes requirements of this document. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies.
GB/T 14926.42 Laboratory animal – Bacteriological examination – Collection of specimens
GB/T 14926.50 Laboratory animal – Enzyme-linked immunosorbent assay
GB/T 14926.51 Laboratory animal – Immunoenzyme assay
GB/T 14926.52 Laboratory animal – Immunofluorescence assay
3 Terms and Definitions
No terms and definitions are required for this document.
4 Principle
4.1 Principle of Antibody Detection
Based on immunological principles, MVM antigen is used to detect MVM antibodies in mouse serum.
4.2 Principle of Nucleic Acid Detection
Based on molecular biology principles, the unique genomic nucleic acid sequences of MVM can be identified by nucleic acid amplification techniques such as polymerase chain reaction (PCR).
5 Reagents and Equipment
5.1 Reagents
5.1.1 Enzyme-linked Immunosorbent Assay (ELISA) Antigen
5.1.1.1 Specific Antigen
Inoculate MVM into susceptible cells [such as mouse embryo (ME) cells, 3T3 cells, etc.] and culture for 7 to 10 days. Harvest when cytopathic effect (CPE) reaches +++ to ++++. Freeze-thaw three times or sonicate. Remove cell debris by low-speed centrifugation. Concentrate the supernatant by ultracentrifugation to prepare the ELISA antigen.
5.1.1.2 Normal Antigen
Supernatant obtained by freeze-thawing and disrupting uninfected normal cultured cells (such as ME cells, 3T3 cells, etc.), followed by low-speed centrifugation to remove cell debris.
5.1.2 Antigen Slides
Inoculate MVM into susceptible cells [such as rat embryo (RE) cells, ME cells, etc.] and culture for 7 to 10 days. When CPE reaches ++ to +++, disperse the cells with trypsin, wash with phosphate-buffered saline (PBS), and prepare smears. Air dry at room temperature. Fix with cold acetone for 10 minutes. Store at -20°C.
5.1.3 Positive Serum
Antiserum obtained by immunizing specific pathogen free (SPF) mice with MVM antigen.
5.1.4 Negative Serum
Serum from SPF mice.
5.1.5 Enzyme Conjugate
Horseradish peroxidase-labeled goat or rabbit anti-mouse IgG antibody; or Horseradish peroxidase-labeled Staphylococcal Protein A (SPA).
5.1.6 Fluorescent Conjugate
Fluorescein isothiocyanate-labeled goat or rabbit anti-mouse IgG antibody.
5.1.7 Nucleic Acid Extraction Reagents
Commercially available nucleic acid extraction kit.
5.1.8 Real-time Quantitative PCR Reaction Mix
Commercially available probe-based real-time PCR master mix.
5.1.9 Negative Control
A sample not containing MVM DNA (can be normal animal tissue or normal culture).
5.1.10 Positive Control
A sample of tissue or cell culture with a defined origin containing MVM.
5.1.11 Primers and Probes
Synthesize primers and probes according to the sequences in Table 1. Prepare primers and probes as 10 μmol/L stock solutions using sterile deionized water. Store at -20°C or below.
5.2 Equipment
5.2.1 Microplate reader.
5.2.2 Fluorescence microscope.
5.2.3 Ordinary light microscope.
5.2.4 37°C incubator or water bath.
5.2.5 Real-time quantitative PCR instrument.
5.2.6 Vortex mixer.
5.2.7 Biosafety cabinet.
5.2.8 Homogenizer.
5.2.9 Centrifuge.
5.2.10 Other equipment: Micropipettes, centrifuge tubes, polystyrene microplates, PCR tubes/plates, etc.
6 Detection Methods
6.1 Sample Collection and Processing
6.1.1 Collect serum according to GB/T 14926.42.
6.1.2 Collect organ tissues, cecal contents or feces, experimental inocula, and environmental samples from laboratory animal facilities according to Appendix A.
6.2 Sample Detection Methods
6.2.1 Use the ELISA method for serological detection, operating according to GB/T 14926.50.
6.2.2 Use the immunoenzyme assay (IEA) method for serological detection, operating according to GB/T 14926.51.
6.2.3 Use the indirect immunofluorescence assay (IFA) method for serological detection, operating according to GB/T 14926.52.
6.2.4 Use the real-time quantitative PCR method for MVM nucleic acid detection, operating according to Appendix A.
7 Result Interpretation
7.1 Interpretation of Antibody Detection Results
For positive results, retest using the same method or another method. If still positive, the result is interpreted as positive.
7.2 Interpretation of Nucleic Acid Detection Results
For positive or suspicious results, retest using the same method. If still positive or suspicious, the result is interpreted as positive.
8 Result Reporting
Issue a report based on the interpreted results.
