Hepatitis B virus serum sample preserving diluent and preparation method thereof

A hepatitis B virus and diluent technology, applied in biochemical equipment and methods, microorganism-based methods, and microbial determination/inspection, etc., can solve the problems of easy bleeding and degradation, poor preservation effect, etc. The effect of reducing and inhibiting DNase activity

CN108998569AInactive Publication Date: 2018-12-14AUTOBIO DIAGNOSTICS CO LTD
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Publication Date
2018-12-14
Estimated Expiration
Not applicable · inactive patent

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Abstract

The invention discloses a hepatitis B virus serum sample preserving diluent. The hepatitis B virus serum sample preserving diluent comprises Trise-HCL, NaCL, BSA, gentamicin sulfate, P-300, sunset yellow, lemon yellow and deionized purified water. On the basis of 1 L of the deionized purified water, the contents of Trise-HCL, NaCL, BSA, gentamicin sulfate, P-300, sunset yellow and lemon yellow are0.01 mol, 8.8 g, 20 g, 0.68 ml, 2.0 ml, 0.01 g, and 0.01 g, respectively. The hepatitis B virus serum sample preserving diluent of the invention has the advantages that the diluent comprises few components; the preparation method is simple; the prepared finished preserving diluent can protect the structure of a viral nucleic acid material from damage, inhibit the activity of DNase and effectivelyprotect the DNAs of a virus from degradation, has the advantages of stability, easy storage, convenience, etc., and is applicable to preservation and dilution of the hepatitis B virus (HBV DNA) serumsample. Test results prove that after addition of the preserving diluent prepared by the invention into the hepatitis B virus (HBV DNA) serum sample, DNAs and RNAs can be completely stored at 37 DEGC for 1 week, stored at 20-25 DEG C for 1 month, stored at 4 DEG C for 6 months and store at -20 DEG C or -80 DEG C for a long time.
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Description

technical field

[0001] The invention relates to biological product technology, in particular to a hepatitis B virus serum sample preservation diluent, and the invention also relates to a preparation method of the preservation diluent. Background technique

[0002] Hepatitis B serum dilution samples are mostly used for kit quality control and third-party quality control. Hepatitis B virus, referred to as hepatitis B virus, is a dsDNA virus belonging to the family Hepadnaviridae. The genome (DNA) of hepatitis B virus is a circular structure surrounded by two helical DNA strands. One of the longer negative strands has formed a complete circle; the other, the shorter positive strand, is semi-circular. Substances that cause DNA degradation include water and nucleases that digest DNA. These two substances are ubiquitous in nature. There are many factors that cause DNA degradation, such as external physical factors, chemical factors, biological factors and so on.

[0003] Conv...

Examples

Embodiment 1

[0024] Example 1 Preparation of Hepatitis B Virus Serum Sample Preservation Diluent

[0025] The first step is to measure an appropriate amount of deionized purified water;

[0026] In the second step, add accurately measured Trise-HCL 0.01mol / L, NaCL 8.8g, BSA 20g, gentamicin sulfate 0.68ml, P-300 2.0ml, sunset yellow 0.01g, tartrazine 0.01g into the deionization purification water and stir well;

[0027] The third step is to adjust the pH of the solution to 7.8~8.2, then add deionized purified water to make the volume to 1L;

[0028] The fourth step is to seal and autoclave (121°C, 15min), then store at 2-8°C.

Embodiment 2

[0029] Example 2 Dilute clinical sample experiment

[0030] 1. Instruments, reagents and samples

[0031] Main instruments: dry constant temperature metal bath, FLEX extractor, biological safety cabinet, pipette, Hongshi real-time fluorescence quantitative PCR instrument, etc.

[0032] Main reagents: Hepatitis B virus nucleic acid assay kit (PCR-fluorescent probe method) from Sun Yat-Sen University Daan Gene Co., Ltd.

[0033] Sample: fresh HBV clinically positive sample.

[0034] 2. Implementation plan

[0035] The virus serum sample preservation diluent prepared in Example 1 is used to dilute the HBV positive sample 5 times, 50 times, 500 times, 5000 times. Two dilutions of the control combination experimental group were designed, and the samples diluted in two different dilutions were extracted and amplified with the hepatitis B virus nucleic acid assay kit (PCR-fluorescent probe method) of Sun Yat-Sen University Daan Gene Co., Ltd.

[0036] Control group: negative seru...

Embodiment 3

[0043] Embodiment 3 storage stability test

[0044] 1. Instruments, reagents and samples

[0045] Main instruments: dry constant temperature metal bath, FLEX extractor, biological safety cabinet, pipette, Hongshi real-time fluorescence quantitative PCR instrument, etc.

[0046] Main Reagent: Hepatitis B Virus Nucleic Acid Detection Kit (PCR-fluorescent probe method) from Sun Yat-Sen University Daan Gene Co., Ltd.

[0047] Sample: fresh HCV pseudovirus sample

[0048] 2. Experimental plan

[0049] Dilute the HBV positive samples 100 times and 1000 times with the hepatitis B virus (HBV DNA) serum sample storage diluent prepared in Example 1, and use them as medium and low concentration test samples, which are marked as P1 and P2 respectively. Place them in -20±5°C environment, 2-8°C environment, room temperature (20-25°C) environment and 37°C environment respectively. The design of this experiment is as follows: mark the diluted samples and place them in the In the specified...