> ---------- > From: Wayne A. C. Harris > Sent: Saturday, 19 January 2002 9:04 > To: Cytometry Mailing List > Subject: Here I am at the alter of FLOW > Wayne, Try adding 0.2% Crystal Violet (in PBS) to reduce autofluorescence of macrophages. Mix with cells then immediately wash off in PBS. An alternative is to gate out highly autofluorescent cells in FL3 or another channel not being used (but you may be gating out cells of interest). Greg Hodge > . > Hello Everyone, > > I've been following this list off and on for a while now and I really > appreciate the fact that so many are willing to share their expertise with > neophytes like myself. Now it is my turn to ask for help. > I am currently doing some work on macrophages trying my darndest to see > them > ingest some GFP expressing bacteria. The problem is that I am working with > these cells straight of a mouse and they produce so much autofluorescence > that it is difficult to distinguish signal from noise at lower > fluorescence > intensities. My question is whether or not it is acceptable and most > importantly believable to remove the noise by subtracting out the > background > using histogram subtraction in order to better see what my 'real' signal > is? > Better yet, has anyone a technique for reducing macrophage > autofluorescence? > I know this question has come up before but I never saw an answer on the > list so thought I would ask first. > > thanks in advance, > > -- > Wayne Harris > Lead Research Specialist > > Department of Microbiology and Immunology > Emory University , School of Medicine > > > >
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