Discussion on How to Improve Data Productivity by Avoiding Cell Line Contamination

What is Cell Line Contamination?

Scientists consider the cell line is contaminated while the cell line genotype is different from its original donor. So for determining whether the cell line is contaminated usually starting from the genotype. The contaminated cell lines can be classified into the false cell line (Eukaryotic cell contamination occurs in the early stage of the cell line, the original cells have ceased to exist, which has completely become another cell line), and the misidentified cell line ( Eukaryotic cell contamination occurs after the cell line is established, and the original cells and newly mixed cells are present in the cell line).

What will Cause Cell Line Contamination?

Cell Line nature enemies include Bacteria, viruses, mycoplasma, yeast and other eukaryotic cells. in the culture procedure, what we offered below will lead to cell line contamination:

A.Confusion of the Mark, which refers to paste the wrong label in the daily culture and the management of the cells.

B.Cross Contamination of Cells, the cell line is contaminated by other cells due to cell reagent utilities, reusable or simultaneous operation of a variety of cells and other reasons. If exogenous cells have a faster propagation rate than the original cell line, exogenous cells will gradually replace the original cell line.

C.Cell culture environment change will lead to cell line contamination. Besides, bacteria, mycoplasma contamination or drug treatment may also change the type of cells.

Why it is important to avoid cell line contaminations

Cell line contamination poses a serious threat to the integrity of biomedical research. Following best practices for cell culture validation, storage and processing can help address this ongoing challenge, thereby improving the reliability of experimental research data.

How to Avoid Cell Line Contamination - Mycoplasma detection

1.Mycoplasma culture, which is regarded as a standard method of detection. This is the most sensitive method, depending on the standard agar plate on the selective and highly enriched growth medium. The appearance will be unique ‘Omelette’, the positive results with this method can prove the mycoplasma contamination. However, this method cannot detect all the species, such as Mycoplasma hyorhinis, DNA and PCR analysis is needed to ensure there is no contamination happened!

2.DNA Staining: DNA staining relies on the Hoechst 33258 stain, which causes DNA-rich nuclei and any mycoplasma in the cytoplasm to fluoresce. The use of this method may lead to false positive for the contamination due to Cell Apoptosis or debris, or false negatives as this is the least sensitive method.

1. PCR: PCR-based mycoplasma detection can be very sensitive and can detect up to 20 copy copies of mycoplasma genomes in 2?L of sample. Mycoplasma detection was achieved by amplifying the highly conserved 16S rRNA operon coding region of the mycoplasma genome.
2. Cell-Line Authentication. ICLAC, which co-built by ECACC, representatives from other international cell culture organizations and respectable scientist commit to supporting the importance of cell line authentication, creating a searchable database of cross-contaminated and misidentified cell lines. This can be used to detect cell line contaminations with the regularly updated database.

5.short tandem repeat str profile. With the ASN-0002-2011 standard method, which analyzed 13 specific sites of STR and several additional sites to determine the integrity of human cell lines authenticity.

Reasonable storage and processing method will decrease the cell line contamination:Proper storage and handling of cells and cell culture reagents, as well as good sterile cell culture techniques, can minimize contaminations and improve reproducibility of experimental data.

These are some details about cell line contaminations and methods can be used for avoiding it!

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