Dear Colleagues, I think that Chemical Cytometry can be an alternative to flow cytometry for those in the cytometry community who have to deal with very few cells (<1000). Chemical Cytometry is our term to describe the multi-component chemical analysis of single cells using the separation and detection tools of instrumental analytical chemistry. We are using Capillary Electrophoresis (CE) with Laser-Induced Fluorescence (LIF) detection for this purpose. In essence in chemical cytometry, a single cell is injected into a separation capillary, lysed inside the capillary and the cellular contents are then separated using CE and monitored at the opposite end of the capillary using an LIF detector. We described the concept of Chemical Cytometry and a prototype instrument in Anal. Chem. 2000, 72, 872-877 (ask me to send a pdf file if interested). Introducing the separation step allows for a multi-component analysis of the cellular contents (theoretically hundreds of components). In addition the technique is characterized by a very decent sensitivity - LIF routinly provides the detection limits of ~100 molecules. Thus, the amount of information one can obtain on a single cell using Chemical Cytometry is considerably superior to that in flow cytometry. Another advantage of Chemical Cytometry is its easy coupling with fluorescence image cytometry to monitor cell's morphology or physiological status prior to the Chemical Cytometry analysis. I have recently used this approach to correlate the cell cycle with a number of enzymatic activities. Such a combination allows to recognize the subpopulations of cells with unique metabolic patterns within the same phase of the cell cycle (see Cytometry 1999, 37, 14-20). Of course, CE is associated with time consumption incomparable with that in flow cytometry (although the amount of information gathered may compensate completely this limitation). I am currently proposing to develop a multi-capillary Chemical Cytometry instrument (think of the PE Applied Biosystems 96-capillary DNA sequencers being used by Celera for human genome project) to considerably increase the throughput. If anybody interested in using such an approach I would be happy to help. I would also appreciate any kind of feedback from the cytometry community. Sergey bunny wrote: > Fellow Flowers: > I wonder if I could gather some opinions on releveance as it relates to > minimum numbers of events collected. > I am assisting a lab in developing FACS analysis of PBMC's and monocytes > in CSF. They informed me that when they run FACS, they are "lucky" to > get 100 events. I raised the questions of relevance, stating that those > 100 "events" could as easily contain some non-cell events as cell > events. They insist their data is good becasue it compares somewhat (?) > with a lab in Europe they are collaborating with. We have had many > conversations on this. > (I should mention- neither lab has direct FACS experience. They got all > their protocols second hand, and just collect what they collect). > Additional information: they are looking at chemokine expression > (another difficult task) on these few cells- and I believe they are > using the PBMC's to set up isotype controls. I doubt they even use comp controls. > > Please throw your 2cents my way. And if any of you do this type of > analysis (few cells) with alternative techniques- I'd love to hear about it! > Thanks- > -- > > Bunny > > ******************************************************* > Bunny Cotleur +*+ Bunny Cotleur > Cleveland Clinic Foundation *+* 2001 Lester RD > Neurosciences NC30 +*+ Valley City, OH 44280 > 9500 Euclid Avenue *+* 330-483-4800 > Cleveland, OH 44195 +*+ bunny@cotleur.com > 216-444-1164 *+* > cotleua@ccf.org +*+ > > ******************************************************* > When you do something, you should burn yourself completely, like a good > bonfire, leaving no trace of yourself. > (Shunryu Suzuki) -- Sergey Krylov, Ph.D. Department of Chemistry University of Alberta Edmonton, Alberta T6G 2G2 Canada Phone: 780-492-9251 Fax: 780-492-8231 E-mail: Sergey.Krylov@UAlberta.Ca Web: http://www.chem.yorku.ca/profs/krylov/
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