A critical evaluation of the magnetic cell sorter and its use in the positive and negative selection of CD45RO+ cells.
In this paper, we report on our year-long experience with the magnetic cell sorter (MACS), and present a critical evaluation of its pitfalls and benefits. Satisfactory separation of lymphocytes into subsets with preservation of function can be achieved, but there are several drawbacks: in comparison with Dynal beads, MACS results in a higher cell loss due to the increased number of separation steps and requires depletion of plastic-adherent cells as these will engulf microbeads and contaminate the enriched fraction, and is more expensive. The advantage of MACS over Dynal beads is that the microbeads are biodegradable and do not interfere with proliferation assays: both the depleted and enriched fractions of cells can therefore be used in culture immediately following separation. We used MACS for the positive and negative selection of CD45RO cells: the enriched fraction was of high purity (greater than 98%), but a depleted fraction of greater than 90% purity could not be obtained even after running the same sample over three separating columns. Dynabeads, on the other hand, achieved 98% pure CD45RO-depleted fractions after three separation runs.
Manyonda
I T
IT
Department of Cellular and Molecular Sciences, St George's Hospital Medical School, London, UK.
Soltys
A J
AJ
Hay
F C
FC
D015703:Antigens, CD / Q000032:analysis*; D015704:CD4 Antigens / Q000502:physiology; D016827:CD8 Antigens / Q000502:physiology; D002469:Cell Separation / Q000379:methods*; D003208:Concanavalin A / Q000494:pharmacology; D005069:Evaluation Studies as Topic; D005434:Flow Cytometry / Q000295:instrumentation*; D006649:Histocompatibility Antigens / Q000032:analysis*; D006801:Humans; D017493:Leukocyte Common Antigens; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology; D007959:Lymphocyte Culture Test, Mixed; D016131:Lymphocyte Subsets / Q000276:immunology; D010835:Phytohemagglutinins; D015203:Reproducibility of Results
D015703:Antigens, CD; D015704:CD4 Antigens; D016827:CD8 Antigens; D006649:Histocompatibility Antigens; D010835:Phytohemagglutinins; D003208:Concanavalin A; D017493:Leukocyte Common Antigens