FYI, Del > ---------- > From: Doug Redelman > Sent: Thursday, September 27, 2001 1:39 PM > To: dyoung@nitromed.com > Subject: Re: 4 color capability of Beckman CoulterEPICS XL > > Hello, > > The 4 color system on the XL works quite well although I cannot > compare it with the FACSCalibur system. The only drawback to the > current system is that one must be particularly careful with color > selection. The system has filters for PMT's to detect ~525nm (FL1, > FITC), ~575nm (FL2, PE), ~620nm (FL3, PE-Texas Red) and ~675nm (FL4, > PerCP or PE-Cy5). The problem is with the PE-TxR conjugates. There is > an inherent difficulty with the tail of the PE emission and some PE-TxR > tandem conjugates may be particularly troublesome. If one selects the > combinations such that the PE-TxR labeled antibody identifies cells that > do not bind the PE labeled antibody in the system, then things can be > fine. It would be worthwhile to try several different sources for > PE-TxR conjugates and check them for possible free PE which of course > only makes things worse. > > There is a better solution, namely to get the appropriate filters to > make FL3 detect ~675nm and FL4 ~>750nm. I have the filter set from > Omega for this purpose and it does work. Coulter is working on a > similar set that also includes a different laser-blocking filter to > improve the passage of the longer wavelengths. There has been > discussion here previously about selecting PMT's to work with longer > wavelengths. I don't know that the Coulter PMT's are optimal, but one > can certainly get decent signals. The most important thing is to check > the antibody conjugates. I have looked at various PE-Cy7 conjugates and > have found substantial differences. > > This year, we have run hundreds of samples on an XL-MCL using the > standard 4 color configuration. I have no complaints with the hardware > or the color compensation circuits at all. There have been problems > relating to spectral overlap and possibly to some conjugates that may > leave something to be desired. I do plan to switch to the system with > PE-Cy7 and expect that it will work very well. > > Doug > -- > ******************************************* > * Doug Redelman, Ph.D. Sierra Cytometry * > * 3150 Susileen Drive Reno, NV 89509 * > * Phone: (775) 329-9772 * > * Email: redelman@concentric.net * > * UNR Cytometry Center (775) 784-6459 * > ******************************************* >
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