SN/T 2497.2-2010Test method of import and export dangerous chemicals. Part 2: Plaque forming cell assay (English PDF)

Foreword

SN/T 2497 "Test Methods for Chemical Safety & Exports dangerous" series of standards is divided into 29 parts.

--- Part 1. mammalian liver cells in vivo DNA synthesis (UDS) test;

--- Part 2. plaque-forming cells (PFC) test;

--- Part 3. Large Zhuan reproduction test;

--- Part 4. Saccharomyces cerevisiae mitotic recombination test;

--- Part 5. testicular cells UDS test;

--- Part 6. mammalian cells in vitro sister chromatid exchange test;

--- Part 7. mouse ear swelling test;

Part --- Article 8. fossa lymph node assay;

--- Part 9. Determination of serum hemolysin test;

Part --- Article 10. T lymphocyte proliferation assay test;

--- Part 11. germline mutation assay;

--- Part 12. single cell gel electrophoresis test;

--- Part 13. fluorescence in situ hybridization;

Part --- Article 14. SDS- polyacrylamide gel electrophoresis;

--- Part 15. PCR-SSCP experiment;

Part --- Article 16. Western-Blot test;

--- Part 17. Behavior of mammalian toxicology tests;

--- Part 18. Detection of DNA adducts;

--- Part 19. NorthernBlot experiment;

--- Part 20. Bradford protein assay;

--- Part 21. agarose gel electrophoresis;

--- Part 22. Tm value determination method of DNA;

--- Part 23. organelle separation experimental methods;

--- Part 24. cellular immunity in vitro assay method;

--- Part 25. humoral immune function tests;

--- Part 26. macrophage function tests;

--- Part 27. Flow cytometry of apoptosis;

--- Part 28. Application shuttle plasmid mutations Inspection;

--- Part 29. Biochemical Oxygen Demand (BOD) was measured.

This section SN/T 2497 Part 2.

The partial modification of the use of the Organization for Economic Cooperation and Development (OECD) Chemicals testing guidelines NO. 490 "plaque-forming cell assay", which has Off the technical content of the above-described method is completely the same, in a standard text format according to GB/T 1.1-2000 made editorial changes.

This section proposed and managed by the National Certification and Accreditation Administration Committee.

This section is responsible for drafting unit. People's Republic of China Tianjin Entry-Exit-Entry Inspection and Hunan People's Republic of China Quarantine Bureau.

1 Scope

SN/T 2497 provisions of this part of the test principle and export of dangerous chemicals - Test methods plaque-forming cell assay, the test Materials, test procedures and test results.

This section applies to the import and export of hazardous chemicals is detected by one or repeated contact with the test substance, the test animals humoral immune function Impact.

2 Terms and definitions

The following terms and definitions apply to this section.

In vitro detection and enumeration produce IgM, IgG and other Ig antibody A method of forming a cell, the cell suspension containing plaque with sheep red Mixed cell suspension, which was uniformly suspended in a liquid or semi-solid agar medium, with the participation of complement, on a red background can be See the transparent region that is an anti-sheep erythrocyte IgM plaque-forming cells.

B lymphocytes stimulated by antigens, proliferation and differentiation into plasma cells, plasma cells produce and secrete antigen-specific antibody process.

3 Test principle

Plaque-forming cell assays (referred PFC test), it is to detect and count to produce IgM, IgG and other Ig plaque-forming cells in a way, Since the test has high specificity, screening and strong, does not require special equipment, etc., it could be used as exogenous chemicals on the body member Indicators of immune function effects.

After the sheep erythrocytes (SRBC) in mice immunized with spleen 4d ~ 5d made of a single cell suspension in semi-solid medium with agar gel SRBC were mixed in a thin layer on a plate or slide paved, and incubated at 37 °C, the participation in the complement have to make antibody-secreting lymphocytes sensitized week Wai SRBC hemolysis reaction to form a visible transparent hemolytic area around it, known as hemolytic plaque. This method of detection cells Mainly IgM plaque-forming cells. Secreting anti-sheep red blood cells to detect IgG antibody-forming cells, we need to join the anti-IgG antibodies to Make sheep red blood cells after the addition of complement lysis.

4 Materials and Reagents

--- Hanks solution, SA buffer;

--- Fresh sheep red blood cells;

--- Guinea pig serum;

--- Agarose (freezing point 42 °C);

--- Clean slide;

---test tube;

--- Plates;

--- Stainless steel mesh;