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Erythrocyte lysis buffer and application thereof

A red blood cell lysate and lysate technology are applied in DNA preparation, recombinant DNA technology and other directions to achieve the effects of simple and fast operation, low manufacturing cost and high purity of white blood cells

Inactive Publication Date: 2014-05-14
SANSURE BIOTECH INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to solve the defects of the existing erythrocyte lysate, to provide a human blood leukocyte separation reagent (erythrocyte lysate) and its preparation method which is quick and easy to operate, high in extraction efficiency, safe to use, and low in manufacturing cost. The isolated white blood cells can be used for subsequent nucleic acid extraction

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0013] 1. Red blood cell lysate and its preparation method:

[0014] Weigh 0.1461g of sodium chloride, add 5ml of triton solution, add 28.8256g of glucose, add 0.6075g of trishydroxymethylaminomethane, add sterile water to 500ml, fully mix and dissolve, use hydrogen Sodium or hydrochloric acid solution to adjust the pH to 8.2. Autoclave for 10 minutes.

[0015] 2. Application of erythrocyte lysate:

[0016] The application object of red blood cell lysate is 100 cases of whole blood samples. The specific operation steps of the application are: add 800 microliters of red blood cell lysate to 200 microliters of whole blood in each case, shake and mix well for 10 seconds to obtain a red transparent and clear mixture; centrifuge the above mixture at a speed of 12000 rpm Minutes, the centrifugation time is 1 minute, the supernatant is removed, and the white precipitate is left. The resulting white precipitate is white blood cells, which can be used for nucleic acid extraction. ...

Embodiment 2

[0020] 1. The erythrocyte lysate and its preparation method in this embodiment are the same as in Embodiment 1.

[0021] 2. Application of erythrocyte lysate:

[0022] The application object of erythrocyte lysate is to contain Epstein-Barr virus DNA concentration of 1×10 4 IU of whole blood samples. The specific operation steps of the application are: add 800 microliters of red blood cell lysate to 200 microliters of whole blood, shake and mix well for 10 seconds to obtain a red transparent and clear mixture; centrifuge the above mixture at a speed of 12000 rpm, Centrifuge for 1 minute, remove the supernatant, and leave a white precipitate. The resulting white precipitate is white blood cells, which can be used for nucleic acid extraction.

[0023] 3. Detection and evaluation of the application effect of red blood cell lysate:

[0024] A commercially available EB virus nucleic acid quantitative detection kit was used to extract and detect nucleic acid from the above leukoc...

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PUM

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Abstract

The invention provides an erythrocyte lysis buffer. The erythrocyte lysis buffer comprises the following components: 4.5-5.5 mmol / L of sodium chloride, 0.8-1.3% (v / v) of triton 100, 300-340mmol / L of glucose and 0.008-0.012 mol / L of trishydroxymethyl aminomethane. The erythrocyte lysis buffer can effectively lyse the erythrocyte in a whole blood sample, separate and enrich leucocyte in the blood, and facilitate the subsequent nucleic acid extraction operation of the leucocyte, so as to obtain the leucocyte DNA in the whole blood sample and the DNA of virus in the leucocyte.

Description

technical field [0001] The invention relates to a red blood cell lysate and its application. Background technique [0002] Red blood cells, also known as red blood cells, are often abbreviated as RBC in routine laboratory tests. They are the most numerous blood cells in the blood. The average number of red blood cells per microliter of blood in normal adults is about 4 million to 5 million for men and 3.5 million for women. ~4.5 million. At the same time, red blood cells are also the most important medium for transporting oxygen through the blood in vertebrates, and they also have immune functions. Mature red blood cells in mammals are anucleate, which means they have lost their DNA. However, hemoglobin, whose content in red blood cells is about 120-150g / L for men and about 105-135g / L for women, has obvious inhibitory effect on PCR amplification. Therefore, when nucleic acid is extracted from a blood sample, it is first necessary to remove red blood cells in the blood sam...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
Inventor 戴立忠唐瑶吴康
Owner SANSURE BIOTECH INC
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