At the NFCR, Larry Sklar and I have spent a lot of time making kinetic measurements using flow cytometry. To measure the response of a cell to a stimulus, in addition to rapidly delivering sample to the cytometer, one must also demonstrate the reagents were combined quantitatively (so you know the concentrations of all species) and mixed completely. With respect to rapid kinetics, the following generalization can be made: If you use a stir bar to mix reagents, you will never get the dead-time below several seconds. Which is not to say you can not measure kinetics at ealier times, it is just that the kinetics you measure will be a combination of mixing and reaction kinetics. The approach we have found most useful is adapted from stopped flow kinetic measurements in enzymology, for example, where reagents are mixed in a mixing tee. Syringe-based sample delivery systems enable quantitative reagent proportioning, mixing, and delivery in under half a second (Cytometry 21:223-9). In principle, flow injection technology should allow similar performance. Setting up such a system is not a trivial undertaking (in terms of money or time), but if one has a commitment to making kinetic measurments it is well worth it. Alternately, the Rapid Kinetic Flow Cytometer at the NFCR is available for use in collaborative projects. Proposals for its use may be sent to me or Larry Sklar. John Nolan John P. Nolan, PhD National Flow Cytometry Resource Life Sceinces Division, LS-5, M-888 Los Alamos National Laboratory Los Alamos, NM 87545 505-667-1623 505-665-3024 FAX nolan@telomere.lanl.gov
This archive was generated by hypermail 2b29 : Wed Apr 03 2002 - 11:49:32 EST