>Dear experts in flow cytometry, > >I have a possibility to use 6 colors FACS. The trouble is that I am >not sure it is possible with the filters I have. >I would very much appreciate any help. >The instrument has three lasers: 325, 488 and 633 nm and I can use >following filters: >380/LP >400/40 >424/44 >500/11 >510/20 >530/28 >575/15 >610/20 >660/13 >670/LP >682/13 >Is it possible to find 6 different flourochromes, which would be >efficiently differentiated with these filters/lasers ? There are no useful fluorochromes for phenotyping that are excited by 325 nm. So you are limited to the 488 and 633 excitation. In addition to bandpass detection filters, it is also important to optimize the beamsplitters and the detectors, especially for measuring the red emitting dyes. Six dyes that would be useful for your system are: Using 488 nm excitation: FITC PE PE-TR (aka ECD from Beckman-Coulter) PE-Cy7 and using the 633 nm excitation: APC APC-Cy7 We use these regularly on our Vantage SE for 6 color work. However, without knowing your instrument configuration I could not say whether they would work for you. I would guess that from the filters you described above, you would have trouble detecting PE-Cy7, APC-Cy7, and quite possibly APC. Marty > >Thank you very much for your help. > >Michal > >-- >Michal Abel, MD, PhD >Laboratory of Clinical Immunology >INSERM U25 >Necker Hospital, 161 Rue de Sèvres. Paris >tel: 33 1 42 19 28 87 >fax: 33 1 44 49 53 74 -- Marty Bigos Director, Flow Cytometry Core Laboratory Gladstone Institute of Virology and Immunology Mail: Box 419100 San Francisco, CA 94141 Packages: 365 Vermont Street San Francisco, CA 94103 Direct Delivery (FedEX, UPS): Gladstone Institute San Francisco General Hospital Building 3 Rm 509 1001 Potrero Avenue San Francisco, CA 94110 (tel) 415-695-3832 (fax) 415-826-8449 mbigos@gladstone.ucsf.edu http://gladstone.ucsf.edu
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