>From: "Chris Worth" <caw@bcc.louisville.edu> >To: Cytometry Mailing List <cytometry@flowcyt.cyto.purdue.edu> >Date: Thu, 22 Apr 99 12:47:20 >Reply-To: "Chris Worth" <caw@bcc.louisville.edu> >Subject: Swapping lasers > > > > >We've got a FacStar Plus with a water cooled laser. The unit is NOT >assembled. We are considering getting it online with an upgrade from the >folks at Cytomation. However, I really need to put an aircooled laser on >this unit. A renovation for water would be a major undertaking. Would it >be possible to do this. Would I need to swap out tips? I figure somebody >out there has done this before. I have zero experience with the FacStar. >if it can't be done, what is the situation these days with chillers? >I haven't been around a chiller in over 10 years. I imagine that > >they've changed a bit too. > >so in a nutshell, can I make the switch to an aircooled laser. if not who makes >a good quiet chiller? > >thanks all, > > >Chris Hi Chris, You will have no problems with an air cooled laser on your FACStar+ as long as you have enough mW output. I have run my FACStar+ with an air cooled laser which I found it under a bench. With 10mW output I get a reasonable signal. I would however recommend at least 25mW (best use 40mW) power output for the laser and change or remove the neutral density filter in front of the FSC detector. I would also recommend making sure that all your optics are very clean, especially the laser path prisms and SSC collection lenses. Check the Dichroics and BP filters and ensure that the PMTs are at their sweet point (highest sensitivity) by loosening their mounting screws and twisting them gently to get the best signal (while running beads). Running a FACStar+ with an air cooled laser at 25mW will allow you to use PerCP in your experiments, indeed BD have said you can use up to 50mW before photo bleaching the dye. I have actually detected PerCP on my MoFlo at 120mW, so I reckon it's a time of flight effect. There would be no need to swap the 70um nozzle, I haven't tried any other size but I reckon as long as you have good laser alignment you should have no problems. There are plenty of manufactures out there who will sell you an air cooled laser e.g. Spectra Physics, you could shop around on the web. However make sure that the lasers output mode is TEM00 (Transverse Emission Mode Zero Zero). This laser mode produces an normal distribution power output profile giving a smaller diameter beam with less divergence. What this means that the maximum power is in the centre of the beam and the power decreases as you move away from the centre. There are other modes with give different power profiles e.g. TEM10 which looks like a target i.e.., the highest power in the middle and then the output rapidly decreases as you move further away from the centre only to increases to maximum again at the edge of the beam. This mode would give poor results by variably exiting the fluorochromes at the laser intersect point. It would be made worse by the elliptical optics on the FACStar plus stretching the beam along the beams' x-axis. In summery then, Clean all optics. Carefully align the laser. Maximise PMT signals (sweet point). Get great results. Hope this helps. Andy. Andy Riddell PNAC MRC-LMB Hills Road Cambridge UK tel (0) 1223 402218 fax (0) 1223 412178 email ar3@mrc-lmb.cam.ac.uk
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