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Pretreatment reagent and method for nucleic acid detection whole blood sample

A whole blood sample, pretreatment technology, applied in the direction of biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of inhibition and low sensitivity, achieve suitable for popularization and application, high practical value, improve extraction purity and The effect of efficiency

Active Publication Date: 2020-02-04
AUTOBIO DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] In view of this, the object of the present invention is to provide an easy-to-operate pretreatment reagent and method for nucleic acid detection of whole blood samples, aiming at the problem of inhibition of subsequent PCR amplification and low sensitivity of red blood cells in whole blood samples existing in the prior art, To remove red blood cells from whole blood samples and eliminate inhibition of PCR amplification

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  • Pretreatment reagent and method for nucleic acid detection whole blood sample
  • Pretreatment reagent and method for nucleic acid detection whole blood sample
  • Pretreatment reagent and method for nucleic acid detection whole blood sample

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Experimental program
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Effect test

Embodiment 1

[0036] 1. Blood sample pretreatment reagent for nucleic acid detection, including the following reagent components:

[0037] 1) Red blood cell lysate: NaCl 10mM; Glucose 250mM; NP40 6v / v%; Tris base 0.05M;

[0038] 2) Whole blood sediment suspension: guanidine hydrochloride 0.4g / ml; fatty alcohol polyoxyethylene ether 14v / v%, the balance is water, and Tris adjusts the pH to 4.1;

[0039] 2. Comparison of test results of samples processed by different pretreatment methods

[0040] Take clinically collected positive EBV samples, use the above pretreatment reagents to process the whole blood samples, and compare them in parallel with the common whole blood pretreatment methods. The processed samples are extracted and tested with the exclusive magnetic bead method extraction kit and fluorescent PCR detection kit at the same time.

[0041] The pretreatment method of the present invention:

[0042] 1) Take 0.2-0.4mL whole blood sample, add 1.2mL of the above erythrocyte lysate, vort...

Embodiment 2

[0053] 1. Blood sample pretreatment reagent for nucleic acid detection, including the following reagent components:

[0054] 1) Red blood cell lysate: NaCl 10mM; Glucose 250mM; NP40 6v / v%; Tris base 0.05M;

[0055] 2) Whole blood sediment suspension: guanidine hydrochloride 0.425g / ml; fatty alcohol polyoxyethylene ether 14v / v%, the balance is water, and Tris adjusts the pH to 4.1;

[0056] 2. Comparison of test results of samples processed by different pretreatment methods

[0057] Take clinically collected positive EBV medium positive, weak positive, and detection limit samples, use the above pretreatment reagents to process whole blood samples, and the red blood cell lysate method in the published patent (wherein the red blood cell lysate contains the following components: 4.5-5.5mmol / L of sodium chloride, 0.8-1.2% (v / v) of Triton 100, 300-340mmol / L of glucose and 0.008-0.012mol / L of trishydroxymethylaminomethane) parallel comparison, the processed sample At the same time...

Embodiment 3

[0073] 1. Blood sample pretreatment reagent for nucleic acid detection, including the following reagent components:

[0074] 1) Red blood cell lysate: NaCl 10mM; Glucose 250mM; NP40 6v / v%; Tris base 0.05M;

[0075] 2) Whole blood sediment suspension: guanidine hydrochloride 0.4g / ml; fatty alcohol polyoxyethylene ether 14v / v%, the balance is water, and Tris adjusts the pH to 4.1;

[0076] 2. Use the pretreatment method of the present invention to process the EBV clinical whole blood samples whose value is traceable to WHO, dilute to obtain the detection limit and quantification limit samples, and cooperate with the exclusive magnetic bead method extraction kit and fluorescent PCR detection kit for detection, specifically Steps include:

[0077] 1) Take 0.2-0.4mL whole blood sample, add 1.2mL erythrocyte lysate, vortex and mix well, lyse the erythrocytes for at least 1min, and obtain a clear and transparent solution;

[0078] 2) Centrifuge at 12000rpm / min for 2min, discard the...

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Abstract

The invention belongs to the technical field of biological detection, and discloses a pretreatment reagent and method for a nucleic acid detection whole blood sample. In a clinical nucleic acid detection process of the whole blood sample, the pretreatment reagent for the nucleic acid detection whole blood sample can be used for effectively removing red blood cells in the whole blood sample and eliminating an inhibition effect on PCR amplification. The pretreatment method for the nucleic acid detection whole blood sample can improve the extraction purity and efficiency of nucleic acid in the sample, and can be matched with a paramagnetic particle method nucleic acid extraction reagent and a fluorescent PCR amplification detection technology to detect virus / human genome nucleic acid in the whole blood sample. In a conventional whole blood sample pretreatment process, the pretreatment reagent and method for the nucleic acid detection whole blood sample have relatively high practical valuefor clinical detection of virus nucleic acid of the whole blood sample, and are suitable for popularization and application.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a nucleic acid detection whole blood sample pretreatment reagent and method. Background technique [0002] The main components of blood are plasma, blood cells and genetic material. Blood contains various nutrients, such as inorganic salts, oxygen, and cell metabolites, hormones, enzymes and antibodies. Therefore, blood stores all kinds of information about human health, and the detection of many diseases requires the analysis of blood indicators. [0003] Erythrocytes, also known as red blood cells, are the most numerous blood cells in the blood. They 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. Red blood cells also don't have mitochondria, which release energy by breaking down gluco...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6806
CPCC12Q1/6806C12Q2527/125Y02A50/30
Inventor 张学增王玮李振红杜美鲁清月付光宇吴学炜苗拥军
Owner AUTOBIO DIAGNOSTICS CO LTD
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