high self fluorescence when doing FACS - (Sep/16/2007 )

I recently ran a flow cytometry of my rat alveolar macrophages. Even the cells only got a high background and the reading is about the same as the cells with my fluorecent labelled LPS. Does any one help me figure out the problem??

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Does anyone know how to make the clumped cells as scattered as they can? This has been said might give high auto fluorescent.

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If these are fixed cells, you may want to lower your PFA concentration to reduce autofluorescence. To reduce clumping, vortex vigorously after fixation and filter your cells before you run them.

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These are not fixed cells. I just treat them and then run them through Flow... Thanks for the suggestion about vortexing. Certainly will do it to see if anything improves.

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