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Method for detecting amino acids in cell culture

A technology of cell culture and detection method, applied in the field of chemical analysis and detection, which can solve the problems of complex detection methods and inaccurate determination of multiple components, so as to improve sensitivity and detection limit, realize high-throughput detection, and avoid derivatization operations Effect

Active Publication Date: 2019-02-15
HJB HANGZHOU CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide a method for detecting amino acids in cell cultures, so as to solve the technical problems in the prior art that the detection methods are complex and various components cannot be accurately measured

Method used

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  • Method for detecting amino acids in cell culture
  • Method for detecting amino acids in cell culture
  • Method for detecting amino acids in cell culture

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] The method for detecting amino acids in the cell culture of the present embodiment comprises the following steps:

[0050] (1) Pre-prepared 1mol / L ammonium acetate solution, sample diluent and electrophoresis buffer, specifically as follows:

[0051] Preparation of 1mol / L ammonium acetate solution: Take a 15mL centrifuge tube, accurately weigh 0.77g of ammonium acetate, add 10mL of water, vortex for 30s, and mix well to obtain a 1mol / L ammonium acetate solution for later use;

[0052] Preparation of sample diluent: Take a 15mL centrifuge tube, add 5mL of methanol, 5mL of water, and 0.1mL of 1mol / L ammonium acetate solution, vortex for 30s, and mix well to obtain the sample diluent for later use;

[0053] Preparation of electrophoresis buffer: Add 50mL of mass spectrometry grade methanol, 50mL of mass spectrometry grade water, and 1mL of mass spectrometry grade formic acid into a clean 100mL reagent bottle, and shake well;

[0054] (2) Prepare amino acid standard soluti...

Embodiment 2

[0071] This embodiment refers to the detection method of Example 1, the only difference is that only one test sample is prepared, and the dilution factor is 150 times.

experiment example 1

[0073] In order to compare and illustrate the detection method of the present invention, the detection method of Example 1 and the existing high performance liquid chromatography were used to detect the content of amino acids in the same cell culture, and the test results are shown in Table 4. Among them, the high-performance liquid chromatography technology is the amino acid detection kit of Waters Company.

[0074] Comparison of amino acid detection results in table 4 cell culture

[0075]

[0076]

[0077] It can be seen from the above table that the detection method of the present invention can effectively detect glutamine, tryptophan and cystine, and distinguish glycine and arginine, and can truly reflect the content of cystine in the culture. It solves the technical problem that glutamine, tryptophan and cystine cannot be detected by high performance liquid chromatography, and some amino acids cannot be effectively separated.

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Abstract

The invention relates to the technical field of chemical analysis detection and in particular relates to a method for detecting amino acids in cell cultures. The method for detecting the amino acids in the cell cultures comprises the following steps: carrying out centrifugal treatment on a cell culture to be detected, collecting supernate, and diluting for 100-500 times so as to obtain a test sample; detecting the test sample by using a capillary electrophoresis-mass spectrum method so as to obtain an ion flow chromatogram; acquiring a corresponding peak area of the ion flow chromatogram of amino acids to be detected in the test sample, and substituting into a standard curve of a corresponding amino acid standard product, thereby obtaining the contents of the amino acids to be detected inthe cell culture. Due to certain pretreatment on the cell culture, together with a capillary electrophoresis-mass spectrum technique, and through quantification of an external standard method, the contents of the amino acids in the cell culture can be rapidly detected, the contents of the amino acids in the cell culture can be tested at different stages of cell culture, and consumption of cells upon amino acids can be understood.

Description

technical field [0001] The invention relates to the technical field of chemical analysis and detection, in particular to a method for detecting amino acids in cell culture. Background technique [0002] In the pharmaceutical industry, biomedical technology is booming. Drug proteins depend on cell or microbial expression. For example, in the process of continuous culture and perfusion culture, it is necessary to understand the consumption of amino acids by cells to further optimize the process, so it is very important to be able to determine the amino acid content of the culture. In addition, the medium development process is an important component of the upstream cell culture process, and the detection of amino acid content can also assist in the optimization of the medium process and promote the improvement of the expression of the target protein by cells. Due to the high cost of cell culture media, especially commercial culture media, the ingredients are unknown, and the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/64G01N30/72G01N30/86
CPCG01N30/64G01N30/72G01N30/8675
Inventor 张庆鸿魏昱
Owner HJB HANGZHOU CO LTD