Hi Howard et al, It was Ronald Reagan who attempted to champion the benefits of a small, narrow beam laser, I believe. The mighty Bryte did give quite excellent peak cv's, especially when using a mercury lamp and giving the PI a nice shot of some 550 excitation, but there weren't enough folks that wanted to buy an arc lamp machine for DNA analysis only, and the trade-off you got with FITC excitation, plus the cover slip change and periodic need for manual alignment... . Regards, Ron >Nick Terry writes: > >>IMHO adequate DNA measurements can only be made if the >>cross section of your interogating beam is substantially narrower than the >>diameter of the objects that you are measuring. >> >>(Waiting for flames.) >> >>Under these conditions 2 G1 cells stuck together will have a very similar >>integral (yes - mostly the same as area) signal as a true single G2/M cell. >>The peak signal from a G2/M cell will however be significantly greater than >>that from a G1 cell and from the sequential peak signals from a G1G1 >>doublet. But, the latter is only visible if the exciting beam is narrow. >>In comparisons between area/width and integral (area)/peak discriminating >>ability on an instrument with 5 micron excitation optics the integral peak >>analysis gave the best discrimination. It is even possible to recognize >>doublets that are not progressing linearly through the beam due to >>turbulence or other factors. >> >>How anyone manages to discriminate doublets from true G2 cells with a 40 >>micron beam, or nuclei with a 15-20 micron beam is beyond me. (tactfully). >> >>(Pondering the likelihood that there is a greater proportion of tetraploid >>tumors in the literature than in reality.) > >Yes, one does need a narrow beam for doublet discrimination, but the >most precise DNA measurements are made with arc source systems, >notably the Partec, but also Lindmo and Steen's (Skatron, Bruker, >Bio-Rad, etc.) design, where the beam is substantially larger than >the cells, and the nondirectional illumination is important in >getting the precision... > >Only a warmish breeze; not a flame, I hope. > >-Howard Ronald P. Dudek Clinical Products Business Manager Miltenyi Biotec, Inc. 251 Auburn Ravine Rd. Suite 208 Auburn, CA 95603 Phone (800) 367-6227 Fax (530) 888-8925
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