Freeze-drying protective agent, fluorescent PCR detection kit and freeze-drying process

A technology of freeze-drying protective agent and detection kit, applied in the field of freeze-drying, can solve the problems of large workload, high cost, long cycle, etc., achieve good protection effect, strong versatility, improve freeze-drying activity and storage stability Effect

Active Publication Date: 2022-02-08
无锡百泰克生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to differences in sources, components, proportions and preparation processes of fluorescent PCR reagents produced by various manufacturers, it is necessary to screen suitable lyo

Method used

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  • Freeze-drying protective agent, fluorescent PCR detection kit and freeze-drying process
  • Freeze-drying protective agent, fluorescent PCR detection kit and freeze-drying process
  • Freeze-drying protective agent, fluorescent PCR detection kit and freeze-drying process

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0057] Example 1 lyophilized protective effect on different manufacturers enzyme raw materials

[0058] (1) Preparation 2 × lyophilized protective agent: It is called 8 g of Hyrants 8 g of hydroxyethyl starch, and 100 μl of Tween 20 is added, and the sterile purified water is added to 100 ml, and then use.

[0059] (2) Preparation of fluorescent PCR reagents: Purchase of all reagent manufacturers with low (<1%) of freeze-dried TAQ, reverse transcriptase, RNA enzyme inhibitor three enzyme raw materials for RT-PCR system construction, After adding influenza virus detection primers, probes, etc.

[0060] Among them, each reagent manufacturer is a common fluorescent PCR reagent manufacturer enzyme raw material manufacturer, including Roche Diagnostic Products (Shanghai) Co., Ltd., Nanjing Nuo Weihan Biotechnology Co., Ltd., Meridianbioscience, and Sheng Sheng Biotechnology (Shanghai ) Co., Ltd., Zhuhai Ba Rui Biotechnology Co., Ltd., Wuhan Aibotic Biotechnology Co., Ltd.

[0061] (3) ...

Example Embodiment

[0094] Example 2 lyophilized experiments using a Wellper Bottle

[0095] (1) Preparation of 5 × lyophilized protector: 10 g of Hyrants 40g and hydroxyethyl starch were weighed, and 1.5 ml of Tween 20 was added, and the sterile purified water was added to 100 ml, and for use.

[0096] (2) Preparation of fluorescent PCR reagents: Nanjing Novani Biotechnology Co. , Probe, etc.

[0097] (3) Preparation of a test mixture: The above 5 × lyophilized protective agent is mixed with the fluorescent PCR reagent to obtain a PCR reaction system. Among them, the final concentration of each component of the lyophilized protective agent is shown in Table 8, and the other components are shown in Table 9.

[0098] Table 8 Each component of lyophilization protective agent final concentration

[0099]

[0100] Table 9PCR reactivity in each component concentration

[0101]

[0102]

[0103] The above-mentioned mixed mixture is dispensed with 500 μL to 2 ml of Wolin bottle, and in the cryogenic d...

Example Embodiment

[0120] Example 3 lyophilized experiments using microfluidic chips

[0121] (1) Preparation of 2.5 × lyophilized protective agent: Take the trehalose 5g and hydroxyethyl starch, and 125 ml of Tween 20 is added, and the sterile purified water is added to 100 ml, spare.

[0122] (2) Preparation of fluorescent PCR reagents: PCR system constructed by Zhuhai Ba Rui Biotezen Co., Ltd., reverse transcriptase, RNA enzyme inhibitor three enzyme raw materials, and then add influenza virus detection primers, Probe. The following 8 systems were formulated, in which systems 1 to 6 were fluorescent RT-PCR systems; systems 7 and systems 8 were DNA amplification systems, and reverse transcriptase and RNase inhibitors were not added.

[0123] (3) Preparation of a test mixture: The above 2.5 × lyophilized protective agent was mixed with the fluorescent PCR reagent to obtain a PCR reaction system. Among them, each component of the lyophilizer is shown in Table 14, and other components are shown in Ta...

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PUM

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Abstract

The invention provides a freeze-drying protective additive, a fluorescent PCR detection kit and a freeze-drying process, wherein the freeze-drying protective additive is used for freeze-drying a fluorescent PCR reagent, and the freeze-drying protective additive comprises the following components: trehalose, hydroxyethyl starch and Tween 20. According to the freeze-drying protective agent, the trehalose, the hydroxyethyl starch and the Tween 20 are adopted, so that the active conformation of protein is maintained in the freeze-drying process, the freeze-drying protective agent has a proper glass transition temperature, the protein is well protected in the freezing and dehydration processes, and the inhibition effect on PCR is avoided. The freeze-drying protective agent can be applied to freeze-drying of fluorescent PCR reagents of different manufacturers, and the product is high in universality and has no inhibition effect on the performance of the fluorescent PCR reagents.

Description

technical field [0001] The invention relates to the technical field of freeze-drying, in particular to a freeze-drying protective agent, a fluorescent PCR detection kit and a freeze-drying process thereof. Background technique [0002] During the freeze-drying process of fluorescent PCR reagents, a lyoprotectant must be added to protect the inherent structure of the protein and prevent protein active substances from being damaged during the freezing and drying process. However, due to differences in sources, components, proportions and preparation processes of fluorescent PCR reagents produced by various manufacturers, it is necessary to screen suitable lyoprotectant components and optimize the reaction system for fluorescent PCR reagents produced by different manufacturers. The process cycle is long, the workload is heavy, and the cost is high. Contents of the invention [0003] The main purpose of the present invention is to provide a lyoprotectant, a fluorescent PCR de...

Claims

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Application Information

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IPC IPC(8): C12Q1/686
CPCC12Q1/686C12Q2563/107C12Q2527/125C12Q2565/629
Inventor 蔡禹希李锦锦周志图
Owner 无锡百泰克生物技术有限公司
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