Application and method adopting single sulfonic acid tetrazolium salt to detect cell activity

A monosulfonic acid tetrazolium salt and a detection method technology, applied in the field of medical biological detection, can solve the problems of incomplete dissolution, affecting the accuracy of the experiment, and easy to take away cells, etc., achieving low interference, low cytotoxicity, and wide application range wide effect

Pending Publication Date: 2017-11-21
HANGZHOU JENNIFER BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

MTT (3(4,5-dimethyl-2-thiazolyl)-2 5-phenyl-2-tetrazolium) is a dye that accepts hydrogen ions, which is produced by TTC in the detection of cell viability assays Formazan has very poor water solubility and needs to be dissolved by adding organic reagents (such as DMSO), which is prone to incomplete dissolution and affects the accuracy of the experiment, and it is easy to take away some cells when absorbing the supernatant.
On the other ha

Method used

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  • Application and method adopting single sulfonic acid tetrazolium salt to detect cell activity
  • Application and method adopting single sulfonic acid tetrazolium salt to detect cell activity
  • Application and method adopting single sulfonic acid tetrazolium salt to detect cell activity

Examples

Experimental program
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Effect test

Embodiment 1

[0066] Escherichia coli cell viability detection method, comprises the following steps

[0067] Escherichia coli was inoculated in LB solid medium and cultured overnight in a constant temperature and humidity box at 37°C; expanded in LB liquid medium at a ratio of 1:10 to 1:20, cultured in a shaker at 37°C and 220rpm for 16 to 22 hours; After that, the measured OD 600 3 to 5, inoculate in 96-well plates filled with culture medium, and serially double-dilution. Add monosulfonic acid tetrazolium salt detection reagent and incubate for 0-1 hour, measure absorbance at 450 nm, and evaluate cell viability.

Embodiment 2

[0069] Saccharomyces cerevisiae cell viability detection method, comprises the following steps

[0070] Saccharomyces cerevisiae was inoculated in yeast medium, cultured in a shaker at 30°C, 220rpm for 18-22 hours; after the time was up, measure the OD 600 3 to 5, inoculate in 96-well plates filled with culture medium, and serially double-dilution. Add monosulfonic acid tetrazolium salt detection reagent and incubate for 0-1 hour, measure absorbance at 450 nm, and evaluate cell viability.

Embodiment 3

[0072] Streptomyces cell viability detection method, comprises the following steps

[0073] Streptomyces was inoculated in Streptomyces culture medium, cultured in a shaker at 28°C, 250rpm for 18-22 hours; after the time was up, the OD was measured 600 3 to 5, inoculate in 96-well plates filled with culture medium, and serially double-dilution. Add monosulfonic acid tetrazolium salt detection reagent and incubate for 0-1 hour, measure absorbance at 450 nm, and evaluate cell viability.

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Abstract

The invention relates to the technical field of medical biological detection and provides a cell activity detection method. Color changing of a novel single sulfonic acid tetrazolium salt reagent is utilized to monitor the cell activity in real time. A single sulfonic acid tetrazolium salt detecting system is selected for the testing agent, and the good linear dose relation is taken on the NAD(P)H level and the living cell number. According to the testing agent, through the absorbance value of a single sulfonic acid tetrazolium salt reactant at 450 nm, an EC50 curved line is obtained, and evaluation of the cell activity is conducted. The detecting method is based on the cell high sensitiveness. Single sulfonic acid tetrazolium salt is small in cell toxicity and suitable for a living cell foundation test. The single sulfonic acid tetrazolium salt reagent is mild, and detected cells can be further used for the subsequent test after being rinsed. The single sulfonic acid tetrazolium salt reagent is not prone to reacting with reducing substances, and interference is small. The reagent is wide in application range and is suitable for detection after environmental microorganism enrichment, detection of the activity of algae, fungi and the animal and plant cells and detection of the cells and tissue lesion.

Description

technical field [0001] The invention relates to the technical field of medical biological detection, in particular to the application and method of using monosulfonic acid tetrazolium salt for detecting cell viability. Background technique [0002] NAD(P) + / NAD(P)H is an important coenzyme factor that exists in all organisms, including bacteria, actinomycetes, cyanobacteria, yeasts, molds, fungi, general animals and plants, and participates in various life processes as a reaction medium , including energy metabolism, mitochondrial function, calcium homeostasis, oxidative stress, gene regulation, immune function, aging, and cell death. Therefore, a sensitive and low-toxic detection method based on the cell level to measure changes in NAD(P)H or its related enzyme reactions plays a key role in applications such as scientific research, environmental maintenance, and medical diagnosis. [0003] Several different types of tetrazolium salts have been reported to measure cell vi...

Claims

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Application Information

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IPC IPC(8): C12Q1/04C12Q1/02
CPCC12Q1/04G01N33/5014
Inventor 阮奔放阮健昵福
Owner HANGZHOU JENNIFER BIOTECH CO LTD
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