Method for Quantitative Detection of Dioxins and Screening of Aryl Hydrocarbon Receptor Active Substances Using Reporter Gene Recombinant Cells

Abstract

The invention discloses a method for quantitatively detecting dioxins and screening aromatic hydrocarbon receptor active substances using reporter gene recombinant cells. The method includes: CBG2.8D cells are divided into 4×10 4 The cells / well were inoculated into a 96-well plate and incubated for 24 hours; the sample to be tested was dissolved in DMSO and mixed with the cell culture medium, wherein the volume ratio of DMSO to the cell culture medium was 1:100, and the obtained sample to be tested was cultured The cell culture medium in the 96-well plate was replaced with liquid solution, and the culture was continued for 24 hours; the culture medium was removed, the cells were lysed, and the luciferase luminescence value was detected on the machine, and the content of dioxin-like compounds in the sample to be tested was determined by using the curve. The present invention performs quantitative standard treatment on cells from cultivation to detection, reduces professional requirements for operators, and synchronizes cell culture and cell detection cycle through the control of cell quantity, effectively simplifies operation steps, improves operation timeliness and reduces repeated steps, and saves reagents Use of consumables.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for quantitatively detecting dioxins and screening active substances of aromatic hydrocarbon receptors using reporter gene recombinant cells. Background technique [0002] Dioxin-like compounds (PCDD / Fs and dioxin-like polychlorinated biphenyls DL-PCBs) are typical persistent organic pollutants (Persistent Organic Pollutants, POPs) with high toxicity, persistence, bioaccumulation and long-term Four characteristics of distance mobility. Based on these characteristics, dioxin-like compounds can be widely distributed in various environmental media such as the atmosphere, soil, and water, and can enter the human body through the food chain. Its extremely difficult to degrade and persist, resulting in the continuous accumulation of dioxin-like compounds in the environment and in the human body, eventually posing a threat to human health. The toxic effects of dioxins are mediate...

AI Technical Summary

Problems solved by technology

However, since the system is used for the screening and detection of dioxin-like pollutants, its application range will be widely extended to testing companies, production enterprises and even self-employed units other than scientific research units. The users in these places are not full-time personnel, and some If you do not have the skills of cell culture, if you just follow the general steps of cell culture and do not set the corresponding reference standards in the key steps, then when the cells grow poorly or the cells are not handled properly, the operator will not be able to carry out professional Therefore, it is necessary to form a set of standard cell detection methods to ensure that the system can eliminate the shackles of professional operations in actual work and ensure that cell detection (cell Culture) can perform the detection function smoothly under the guidance of standard steps, and improve the stability and accuracy of cell detection

[0012] In addition, the existing reporter gene detection methods are designed based on laboratory operations, so the detection method steps do not consider the detection efficiency and the cost of reagent consumables in the operation process

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