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L-乳酸作为细胞代谢状态的指示剂:一种简单且经济高效的比色 L-乳酸测定法。

L-lactate as an indicator for cellular metabolic status: An easy and cost-effective colorimetric L-lactate assay.

机构信息

Institute for Immunology, Philipps-University Marburg, BMFZ, Marburg, Germany.

出版信息

PLoS One. 2022 Jul 22;17(7):e0271818. doi: 10.1371/journal.pone.0271818. eCollection 2022.

Abstract

BACKGROUND

In recent times, the study of metabolic pathways has become inevitable and predominant for a variety of research fields as cancer biology and immunology. L-lactate as a product of anaerobic glycolysis has shown to be an important indicator of the cellular metabolic status and can be associated with diverse cellular effects. For this reason, L-lactate assay kits are of high demand when metabolic effects need to be considered. Nevertheless, commercially available kits are not affordable if multiple samples must be evaluated.

PRINCIPAL FINDING

In this work, we develop an easy and cost-effective colorimetric assay for quantification of L-lactate suitable for cells with low or high L-lactate production based on LDH activity and suitable for 96 well-plate format. Using different metabolic regulators, we demonstrate the capacity of the assay to detect and quantify L-lactate from the supernatant of HeLa cancer cell line. Furthermore, we validate the assay against a commercially available kit by demonstrating no significant difference between both assays. Finally, we show that the assay is capable of quantifying L-lactate in primary cells such as hPBMCs that were stimulated with toll-like receptor ligands and treated with different metabolic regulators.

CONCLUSION

We herein present an easy custom assay that is suitable for cells with low and high L-lactate production at very low cost compared to commercially available kits. These advantages of the custom assay can simplify the research in the field of metabolism and related fields.

摘要

背景

近年来,代谢途径的研究已成为癌症生物学和免疫学等多种研究领域不可或缺且主要的研究方向。L-乳酸作为无氧糖酵解的产物,已被证明是细胞代谢状态的一个重要指标,并与多种细胞效应相关。出于这个原因,当需要考虑代谢效应时,L-乳酸检测试剂盒的需求量很大。然而,如果需要评估多个样本,市售试剂盒的价格就难以承受。

主要发现

在这项工作中,我们基于 LDH 活性开发了一种简单且经济高效的比色法 L-乳酸定量测定法,适用于低产或高产 L-乳酸的细胞,适合 96 孔板格式。使用不同的代谢调节剂,我们证明了该测定法能够从 HeLa 癌细胞系上清液中检测和定量 L-乳酸。此外,我们通过证明两种测定法之间没有显著差异,对该测定法进行了商业化试剂盒的验证。最后,我们表明该测定法能够定量检测经 Toll 样受体配体刺激并用不同代谢调节剂处理的原代细胞(如 hPBMCs)中的 L-乳酸。

结论

我们在此提出了一种简单的定制测定法,与市售试剂盒相比,成本非常低,适用于低产和高产 L-乳酸的细胞。这种定制测定法的优势可以简化代谢及相关领域的研究。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/c778/9307176/4de27ca76b84/pone.0271818.g001.jpg

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