Regina hi One of the things that might work would be to use a biotinylated primary antibody and then come back with anti-biotin FITC. The background is in fact quite less as compared to streptavidn or avidin. Vector use to make an anti-biotin FITC. I am not sure if anybody makes anti-Biotin PE. If any body knows I would be interested in finding out. Hope this helps. Joseph Mattapallil Asst. Research Immunologist School of Medicine University of California Davis Davis, CA 95616 At 9:03 AM -0400 8/2/00, Regina Harley wrote: >Regina H. Harley, M.S. >Lab Supervisor - Flow Cytometry Facility >Center for Vaccine Development >University of Maryland - Baltimore >685 West Baltimore Street >Baltimore, MD 21201 >(410) 706-7376 FAX: (410) 706-6205 >rharley@medicine.umaryland.edu > > >>> ttown <ttown@luna.cas.usf.edu> 07/31/00 12:18PM >>> > >The specific question is, do these percentages REALLY mean that out of >100 cells, x percentage express CD40? I tend to think that many more >of >those cells actually express CD40 but that such low expression levels >of >CD40 are undetectable by FACS. If this is the case, then what do >these >percentages really mean, and is there a reference(s) for this? > >Best wishes, > >Terrence Town > > >This actually is a very good question and is relevant to any low >incidence/low percentage molecule (where the incidence is the number of >molecules per cell and the percentage is the number cells with the >molecule). > >We are going to be working with a target that has both a low incidence >and a low percentage. Due to a number of technical issues, the only >fluorochromes we can use are PE and APC (and therefore can't use >Molecualr Probes FITC amp. kit). Does anyone have a working system of >amplification for these fluorochromes? We been trying various multistep >procedures but have been having problems with high background. Any >help/suggestions would be appreciated. > >Thanks very much. > >Regina
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