Peter, et al- I can attest to the fact that viability goes down the tubes with Hoechst 33342 and UV power > 50 mw with 10 usec transit of the laser beam, at least on some common tissue culture cell lines. But the viability issue (cells with compromised membrane integrity not being able to exclude a dye such as DAPI) is the flip side. The dead or compromised cells that take up the dye are the ones you want to exclude anyway and you don't care if they get fried. Robb At 07:01 PM 3/14/2001 -0500, PLopez@adarc.org wrote: >Hi Mark, >That's a good question, and although I've never heard >people complain about viability after a sort with this technique, >I never did the actual post-sort viability study. My experience is >using Hoechst, not DAPI, but in either case I wouldn't be worried. >The sperm sorting folks have looked into this. They do UV excited >Hoechst 33342 sorting daily with good results. >The live/dead app gives a bright staining so you don't need much >UV power. I've had good resolution using a sick argon 90-5 which >only put out 30mw of UV . I bet you could even use half that power. >The viable cells are negative, so they don't really pick up much dye, if >that is the concern. > >Peter Lopez >The Aaron Diamond AIDS Research Center >212.448.5188 (office) >212.448.5159 (fax) >212.448.5190 or 5110 (lab) > > > > "Mark > Kukuruga" To: Cytometry Mailing List > <kukuru@med.u <cytometry@flowcyt.cyto.purdue.edu> > mich.edu> cc: > Subject: DAPI as a viability > exclusion > 03/14/2001 dye > 01:34 PM > > > > > > >Hi all . . . >Considering the use of DAPI as an exclusion dye, I'm wondering about the >effects of >(albeit brief) UV irradiation on the cells that we sort "viably." Should >we worry, >or is it negligible? >MAK. > > >-- >Mark A. KuKuruga, Managing Director >University of Michigan Flow Core >7416 CCGC 0946 >(734) 647-3216, fax (734) 936-7376 >kukuru@umich.edu
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