Recently, I encountered some difficulties while attempting to sort 4T1 cells. The main issue is that 4T1 cells like to stuck together, which has led to a considerably low yield of sorted single cells that fails to fulfill my experimental requirements. I would be extremely grateful if anyone could offer me some suggestions.
I carried out the cell digestion process using a 0.25% trypsin-EDTA solution for ten minutes and then terminated the digestion with 1640 medium supplemented with 10% FBS.
For routine passaging, cells are recommended to be split at a 1:10 ratio every 3-4 days. 4T1 cells frequently clump during growth. When confluent sections of cells become large (e.g. fill an entire field of view at 10X) the cells should be passaged to minimize cell death.
Hi Huang,
- 4T1s are sticky by nature, and long trypsinization (10 min) makes clumping worse — shorten digestion and pipette gently to disperse.
- Pass through a cell strainer before sorting; this improves single-cell yield.
- Keep density moderate — overconfluent cultures tend to aggregate more after dissociation.
- For more uniform suspensions and reduced clumping, DendroTEK dPGA | Protease-Resistant Culture Matrix provides a stable alternative surface that doesn’t degrade like PLO.
Best of luck, Jason Hamlin
Source: I work at DendroTEK Biosciences. www.dendrotek.ca
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