Flowers, I received several helpful replies to my question about methods for flow sorting single cells into wells for pcr amplification. Although there are many good articles, the best general methodological reference that I've seen is: AB Kantor, CE Merrill, and JL Hillson (1996) Construction of cDNA from single unstimulated mouse B lymphocytes: Method and application to the study of expressed antibody repertoires in FACS-sorted murine B cell subsets. Chapter 13 in Weir's Handbook of Experimental Immunology (ed LA Herzenberg), Blackwell Science, Cambridge, MA. pp 13.1 - 13.6 A tip mentioned in this reference, and also reinforced in an e-mail to me from Larry Arnold, is to adjust the voltage to the deflection plates in order to angle the waste stream off to the side and aim the sort stream directly down into pcr tube or well. This increases the likelihood of the sorted cell arriving in the small volume of pcr lysis buffer (and not getting stuck on the side of the tube). Thanks for all your help. Alice Alice L. Givan Englert Cell Analysis Laboratory of the Norris Cotton Cancer Center Dartmouth Medical School Lebanon, New Hampshire NH 03756 tel 603-650-7661 fax 603-650-6130 givan@dartmouth.edu
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