It is a very interesting question. I also observed that camptothecin-treated Jurkat cells had less autofluorescence than the untreated Jurkat cells. Jimin Wang, Ph.D. Biosource International 820 Flynn Road, Camarillo CA 93012 Tel: 805 383-5233 -----Original Message----- From: Andrew D. Wells, Ph.D. [mailto:adwells@mail.MED.UPENN.EDU] Sent: Monday, January 03, 2000 7:13 AM To: cyto-inbox Subject: Re: TUNEL Assay > >Dear Flowers, > >I have a question about the Tunel assay using flow cytometry. I treated my >cells ( adherent cell line) with agents, Actinomysin D, Ceramide, >Campothecin, before staining with dUTP-FITC (kit from boehringer mannhein) >to detect apoptosis.What does mean when the label sample(negative sample) is >higher( histogram peak) than positive sample (TdT sample). >I would appreciate any help anyone can give. Soheila, Although its unlikely that the following point will explain (all) your problem, I thought it would be worthwhile to mention to you and the mailing group that, in my experience, some drugs that one might use to induce apoptosis are retained in the cells and can act as fluorochromes when hit by the laser; i.e., your drug-treated, non-TdT-treated cells can fluoresce quite brightly in FL1 and 2. Again, this can't explain every case of high background in TUNEL, but its important to be cognizant of this situation. And why your TdT-treated would be less fluorescent than your control is not clear to me (quenching of the background by the FITC?). Hope this helps, AW Andrew D. Wells, Ph.D. University of Pennsylvania Department of Medicine 728 Clinical Research Building 415 Curie Boulevard Philadelphia, PA 19104 (215) 573-1840 (office) (215) 898-1951 (lab) (215) 573-2880 (FAX) adwells@mail.med.upenn.edu
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