dust mate
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Need for Help (DC, IL12,LPS, autoMACS)
dust mate replied to Masoud's topic in Microbiology and Immunology
Can not you perform intracellular staining after culture of whole popliteal LN cells instead of purification of DCs? I am not sure even with CD11c MACS you will achieve high purity anyway. these beads work well when initial cell frequency is high (e.g. in the spleen after 2 rounds of AutoMACS I was getting around 60-70% CD11c+ cells (which is low) when initial frequency was 1-3% only. but when initial frequency was around 30% then purity was >97%). -
prepare single cell suspension first and then use no more than 2-3ml buffer per spleen. Don't lyse more than 1 minute! This time is more than enough