Hello everyone,
I recently thawed THP-1 cells, which were frozen at passage 12 in 2016. I got the cells back to culture in RPMI + 10% FCS + 1% HEPES + 1% PenStrep. The cells seemed to recover well after thawing, and I changed the media the day after to get rid of residual DMSO. The cells were cultured in T-75 and then transferred to T-175 flasks.
On day 4, one of the two flasks showed turbid media, which was bright pink (brighter than standard RPMI) and microscopic examination indicated minuscule organisms that were rapidly moving about.
Given that only one of the two flasks showed signs of contamination, we are currently ruling out the possibility of one of the reagents be the source. Although, it might well be a copy number issue.
Please find attached a photo of the cells under the microscope (40x). THP-1 cell is indicated by a black arrow, the red box indicates the contaminant.
In my 8 years of working with various cell lines, I have never seen anything like this!
Any suggestions as to what this might be, and how to get rid of it, would be of great help!
Thank you
Best,
Sripriya
It's really hard to say but it seems to me gram positive cocciea.
I think the best way is culture the media on an enrichment media like BHI.
regards
Sripriya Murthy I don't know what kind of bacteria it is. But I know that your cells were contaminated with bacteria since I ever experienced something like this before. Did any other sample in the same incubator also contaminated before you or at the same time?
Thank you for your answers. Novi Angeline , no there were no other signs of contamination in the other cells that were in culture alongside the THP-1 cells. We suspect it might have been the frozen vial of cells that was the source of contimation.
Sripriya Murthy As other people have suggested it is probably bacteria. When it happened to me I just threw everything away and tried based on the dates I had frozen cells, to identify when the contamination started. If it is a completely out of the blue contamination and it comes back, I would check the filter on the laminar hood. This was the problem for me and lead to a big waste of time trying to determine the origin of the contamination.
Dear Dr Traci L Testerman ,
I found your answer to the expressed question more logical and out of practice. I would like to ask you a question: Is it normal to use flames under the laminar flow to get the mouth of the flasks or other sources of contamination sterile? I have seen that researchers in the same lab have different opinions about that issue. Some researchers believe that using flame under laminar flow will ruin hepa filters and others believe that it is the only way they could get rid of contaminators? I would be thankful to know your opinion.
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