Abstract
To assess the presence and absence of mycoplasma contamination in cell culture, Fourier transform infrared (FTIR) microspectroscopy, coupled with multivariate analysis, was deployed to determine the biomolecular changes in hepatocellular carcinoma cells, HepG2, before and after mycoplasma contamination. The contaminated HepG2 cells were treated with antibiotic BM-Cyclin to decontaminate the mycoplasma, and polymerase chain reaction (PCR) was then performed to confirm the presence or the absence of mycoplasma contamination. The contaminated and decontaminated HepG2 cells were analyzed by FTIR microspectroscopy with principal component analysis (PCA) and peak integral area analysis. The results showed that the FTIR spectra of contaminated HepG2 cells demonstrated the alteration in the IR spectra corresponding to the lipid, protein, and nucleic acid regions. PCA analysis distinguished the spectral differences between the groups of mycoplasma-contaminated and -decontaminated cells. The PCA loading plots suggest that lipid and protein are the main contributed molecules for the difference between these two cell groups. Peak integral area analysis illustrated the increase of lipid and nucleic acid and the decrease of protein contents in the contaminated HepG2 cells. FTIR microspectroscopy is, therefore, proven to be a potential tool for assessing mycoplasma removal by monitoring biomolecular alterations in cell culture.
Highlights
Mycoplasma is a significant issue in research laboratories as contaminants in cell culture.The bacterial contamination potentially leads to disastrous laboratory results since mycoplasma can alter the infected host cell at a molecular level, providing inaccurate and unreliable experimental data.The bacteria compete with the host cell and deplete their nutrients, which disrupt host cell metabolism, morphology, and function at a molecular level
Our study demonstrates the application of Fourier transform infrared (FTIR) microscopy for the cell line inspection of mycoplasma contamination because FTIR spectra reflects the biomolecular alteration
The mycoplasma decontamination was confirmed by the standard polymerase chain reaction (PCR) method
Summary
Mycoplasma is a significant issue in research laboratories as contaminants in cell culture. FTIR spectra from cancerous kidney and ovarian cells could be segregated from their normal counterparts, indicating cellular component alteration toward cancer progression [9,10] These studies have demonstrated the effectiveness of FTIR microspectroscopy to define biomolecular changes during different stages of the cell. Since mycoplasma contamination causes alteration of biochemical composition (i.e., lipid, protein, and nucleic acid) in their host cells [11,12], the detection of overall biomolecular compartments, seems to be useful for examining the effect of mycoplasma contamination in the host cell, as well as discriminating them from decontaminated cells. The obtained data from this study could provide the information on potential FTIR microspectroscopy techniques on the assessment of mycoplasma impact on cellular biomolecules, as well as implying a new mycoplasma detection method in cultured cells
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