Fusion gene indoor quality control product, manufacturing method and recipe of fixing solution of fusion gene indoor quality control product

A technology that integrates genes and indoor quality control. It is applied in the fields of life science and biology and can solve the problems of not being on the market.

Inactive Publication Date: 2016-10-12
合肥艾迪康医学检验实验室有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the relatively late start of domestic related testing, and the target of fusion gene testing is RNA, a nucleic acid that is easily degraded by RNases that are everywhere in the environment, so the quality control substances used to monitor the quality of laboratory testing have not been available on the market.

Method used

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  • Fusion gene indoor quality control product, manufacturing method and recipe of fixing solution of fusion gene indoor quality control product
  • Fusion gene indoor quality control product, manufacturing method and recipe of fixing solution of fusion gene indoor quality control product
  • Fusion gene indoor quality control product, manufacturing method and recipe of fixing solution of fusion gene indoor quality control product

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Extraction of nucleated cells from whole blood and fixation of cell suspension: Add 1ml 1× red blood cell lysate into a 1.5ml centrifuge tube, take 0.5ml of whole blood sample, mix by inverting. Centrifuge at 4000rpm for 3min, discard the supernatant, add erythrocyte lysate to wash once, and add 50ul of fixative solution equal to the volume of the cell pellet, which is the prepared quality control product.

[0031] The fixative includes 4°C pre-cooled absolute ethanol (AR), 70-75ml; glacial acetic acid (AR), 1-5ml; DEPC water to 100ml; refrigerated storage at 4°C.

Embodiment 2

[0033] RNA extraction of the quality control product: take the quality control product in Example 1 and add 1 ml of normal saline, let it stand for 5 minutes, then centrifuge and discard the supernatant. Add 1ml of normal saline to wash again. Add 1 ml Total RNA Isolation Reagent, pipette repeatedly until no obvious cell clumps, add 200 μl of chloroform, vortex and mix for 30 seconds, and let stand on ice for 10 minutes. Centrifuge at 14,000rpm at 4°C for 10min. Use a pipette to draw 450 μl of the supernatant and transfer it to another centrifuge tube, add an equal volume of pre-cooled isopropanol, invert and mix well, and then let stand on ice for 10 minutes. Centrifuge at 14,000rpm at 4°C for 10min. Then wash and centrifuge once with 75% ethanol and absolute ethanol respectively. Dry at room temperature for 5 min, add 50 μl DEPC-H2O to dissolve.

Embodiment 3

[0035] Reverse transcription: Take 4ul of the RNA solution in Example 2 (concentration about 200ng / ul), add 1ul Primer mix (ReverTra Ace qPCR RT Kit) and 3ul DEPC-H2O, mix well, and pre-denature at 70°C for 5 minutes; after quenching on ice for 1 minute, add 5 *RT buffer 4ul (ReverTra Ace qPCR RT Kit), Enzyme Mix 1ul (ReverTra Ace qPCR RT Kit), and add DEPC-H20 7ul to a total volume of 20ul. After reacting at 37°C for 60 minutes, it was inactivated at 98°C for 5 minutes, and the obtained cDNA was the quality control product to be tested.

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Abstract

The invention provides a method of manufacturing a common fusion gene indoor quality control product by using a fixed cell suspension and a recipe of a fixing solution of the fusion gene indoor quality control product. The fusion gene detection capable of being used for indoor quality control comprises four kinds of fusion genes including BCR-ABL P210 (b2a2 type and b3a2 type), BCR-ABL P190, PML-RaRa (L type and S type) and AML1-ETO. The fixing solution used in the method has low price and is non-toxic and harmless; one tube of quality control product can be used for simultaneously completing the indoor quality control of four kinds of fusion genes; convenience and economy are realized; no salable fusion gene quality control products exist on the existing market; the blank of no fusion gene quality control product in the industry at present is filled.

Description

technical field [0001] The invention belongs to the field of life science and biotechnology, and in particular relates to an indoor quality control product for common fusion genes produced by using fixed cell suspension, a production method and the formula of the fixative. Background technique [0002] The incidence of leukemia is increasing day by day, and the incidence rate has almost doubled in the last 10 years. With the development of medical level and technology, the classification, treatment and prognosis evaluation system of leukemia have been systematically improved and perfected. The advent of targeted therapy drugs has made molecular diagnosis an indispensable part of disease diagnosis, treatment and prognosis monitoring. Among them, the detection of fusion genes not only assists in disease classification and diagnosis, but is also closely related to the selection of medication and treatment options. Especially BCR-ABL P210 and BCR-ABL P190 fusion genes are char...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6806C12Q1/6886C12Q2600/112C12Q2600/118C12Q2545/113
Inventor 邹媛陈红梅夏成青
Owner 合肥艾迪康医学检验实验室有限公司
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