Gene, Marcel, and other interested parties, I forwarded your message to John Sharpe, the optics guru here to see if he had any specific information. The message below is what he sent back to me. Hope that is of some help without sounding too corporate. :-) kb --------------- Keith Bahjat, Ph.D. Applications Engineering Manager Cytomation, Inc Fort Collins, Colorado Phone: (970) 226-2200 x223 Fax: (970) 226-0107 keithb@cytomation.com -----Original Message----- From: John Sharpe Sent: Monday, September 24, 2001 9:15 AM To: cyto-inbox Subject: RE: HeCd laser option on LSR Keith, The main difference between the lasers is that the LSR has a Kimmon 325nm 8mW HeCd and the CYAN has 50mW of 351nm from an Enterprise 621 Ar+ laser. There are 3 issues of interest here - i)power, ii)spectral suitability to excitation of a fluorochrome of interest, and iii)optical noise. With regard to i), more power results in better fluorochrome excitation (nearer saturation) and higher signal to noise thus providing a brighter signal (and less pmt amplification). Spectral suitability of the excitation wavelength will of course depend on your fluorochrome, however as an example, Ho33342 has an excitation envelope with a maximum (100%) near 350nm (vs. ~50% at 325nm) thus suggesting the Ar+ line is better suited to this application than the HeCd line. Third, while it is possible to obtain new HeCd lasers with relatively low noise, in my experience these lasers degrade with age reaching optical noise levels in excess of 5% within a few months. To state the optical noise specifications given by the vendors, the HeCd is stated as having <2% RMS noise (30kHz-10Mhz) whilst the Ar+ laser is specified at <1% RMS noise (10Hz to 2Mhz). Keep in mind that it is the cell-to-cell variations were are interested in which is typically in the 0 to 10kHz event per second range. Hope this helps. John -----Original Message----- From: Keith Bahjat Sent: Saturday, September 22, 2001 11:13 AM To: cyto-inbox Subject: RE: HeCd laser option on LSR John, Some users on the cytometry mailing list have questions about a couple of different lasers. Any chance you could clarify things for them? thanks kb -----Original Message----- From: Pizzo,Eugene [mailto:Pizzo@nso1.uchc.edu] Sent: Friday, September 21, 2001 8:51 AM To: cyto-inbox Subject: HeCd laser option on LSR Hi, With the remark I received concerning UV laser reliability in the LSR (see below) I was wondering if anyone could comment on Cytomations claim that their UV laser in their new Cyan machine has less noise and whether the LSR could be retrofitted if need be with a less noisy HeCd. Gene/UCONN Health If you used an LSR be aware that those lasers won't allowed you to performed cell cycle measurements with reasonnable CV for all there life time. In our hands, usually you can perform DNA content and cell cycle for a period of about 8 to 12 months(wich is in accordance with HM Shapiro, Practical flow cytometry).Those lasers became noisy after only a few months and getting worst as time goes by.For calcium flux they are perfect because with Indo-1 you will do a ratio measurement wich eliminated the noise problem. Marcel Manager, Core cytometry lab. INRS-Institut Armand Frappier Université du Québec
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