I am using L6 skeletal muscle cell lines for glucose uptake assay. I am culturing it in DMEM + FBS10% + ANTIBIOTICS1%. Sometimes the cells grow fine with uniform distribution throughout the flask and most of the times they form colonies/clumps. Hence, I never allow the cells to 70-80% confluent. But after sub culturing the cells they form clumps, which does not allow me to seed in to 96 well plate. I never use to agitate or mix the cells forcefully according to ATCC guideline (to prevent forming clumps/colonies).

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