Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

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

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

Active Publication Date: 2022-06-24
BIOTEKE CORP (WUXI) CO LTD
View PDF23 Cites 0 Cited by
  • 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 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

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A freeze-drying protective agent, fluorescent PCR detection kit and freeze-drying process
  • A freeze-drying protective agent, fluorescent PCR detection kit and freeze-drying process
  • A freeze-drying protective agent, fluorescent PCR detection kit and freeze-drying process

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Example 1 Freeze-drying protection of enzyme raw materials from different manufacturers

[0058] (1) Preparation of 2× lyophilized protective agent: Weigh 8 g of trehalose and 8 g of hydroxyethyl starch, add 100 μL of Tween 20, add sterile purified water to dissolve, and dilute to 100 mL for later use.

[0059] (2) Preparation of fluorescent PCR reagents: Purchase lyophilized Taq enzyme, reverse transcriptase, and RNase inhibitor three enzyme raw materials without glycerol or low glycerol content (<1%) from various reagent manufacturers to construct RT-PCR system. Then, primers and probes for influenza virus detection are added.

[0060] Among them, each reagent manufacturer is a common fluorescent PCR reagent manufacturer and enzyme raw material manufacturer on the market, including Roche Diagnostics (Shanghai) Co., Ltd., Takara Bio Inc., Nanjing Novizan Biotechnology Co., Ltd., MeridianBioscience, Yisheng Biotechnology (Shanghai) Co., Ltd. ) Co., Ltd., Zhuhai Baorui ...

Embodiment 2

[0094] Example 2 Freeze-drying experiment using vial as carrier

[0095] (1) Preparation of 5× lyophilized protective agent: Weigh 40 g of trehalose and 10 g of hydroxyethyl starch, add 1.5 mL of Tween 20, add sterile purified water to dissolve, and dilute to 100 mL for later use.

[0096] (2) Preparation of fluorescent PCR reagents: Three kinds of enzyme raw materials, namely Taq enzyme, reverse transcriptase, and RNase inhibitor, which are free from glycerol and can be freeze-dried from Nanjing Novizan Biotechnology Co., Ltd. are used to construct a PCR system, and then primers for influenza virus detection are added. , probes, etc.

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

[0098] Table 8 Final concentration of each...

Embodiment 3

[0120] Example 3 Freeze-drying experiment using microfluidic chip as carrier

[0121] (1) Preparation of 2.5× lyophilized protective agent: Weigh 5 g of trehalose and 10 g of hydroxyethyl starch, add 125 mL of Tween 20, add sterile purified water to dissolve, and dilute to 100 mL for later use.

[0122] (2) Preparation of fluorescent PCR reagents: The three enzyme raw materials of glycerol-free lyophilized Taq enzyme, reverse transcriptase and RNase inhibitor from Zhuhai Baorui Biotechnology Co., Ltd. were used to construct a PCR system, and then primers for influenza virus detection, probe, etc. A total of the following 8 systems were prepared, of which systems 1 to 6 were fluorescent RT-PCR systems; systems 7 and 8 were DNA amplification systems without reverse transcriptase and RNase inhibitors.

[0123] (3) Preparation of test mixture: mixing the above 2.5× lyophilized protective agent and fluorescent PCR reagent to obtain a PCR reaction system. Among them, the final con...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention provides a freeze-drying protectant, a fluorescent PCR detection kit and a freeze-drying process thereof, wherein the freeze-drying protectant is used for freeze-drying fluorescent PCR reagents, and the freeze-drying protectant includes the following components: trehalose, hydroxyethyl base starch and Tween 20. The lyoprotectant of the present invention adopts trehalose, hydroxyethyl starch and Tween 20, which not only maintains the active conformation of the protein and has a suitable glass transition temperature during the freeze-drying process, but also protects the protein during the freezing and dehydration process. To a very good protective effect, and will not inhibit the PCR reaction. The lyoprotectant can be applied to the lyophilization of fluorescent PCR reagents from different manufacturers, and the product has strong versatility and has no inhibitory 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. Background technique [0002] During the freeze-drying process of fluorescent PCR reagents, a freeze-drying protective agent must be added to protect the inherent structure of the protein and prevent the 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 different manufacturers, the fluorescent PCR reagents produced by different manufacturers need to be screened for suitable lyoprotectant components, and the reaction system should be optimized. The process cycle is long, the workload is large, and the cost is high. SUMMARY OF THE INVENTION [0003] The main purpose of the present invention is to provide a freeze-dryi...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/686
CPCC12Q1/686C12Q2563/107C12Q2527/125C12Q2565/629
Inventor 蔡禹希李锦锦周志图
Owner BIOTEKE CORP (WUXI) CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com