Magnetic affinity cell sorting and its biological applications

Magnetic affinity cell sorting (MACS) method is a powerful tool often used to identify and fractionate heterogeneous cell populations into pure populations. Previous methods made use of endogenously expressed surface proteins for separation of cell types, which are limited to cells expressing a surface marker. A novel approach was developed in this study in which cells were rendered to express a surface protein by DNA-mediated transfection. The MACS method was optimized by cotransfecting the plasmids encoding the reporter chloramphenicol acetytransferase (CAT) and either the vesicular stomatitis virus G (VSV-G) protein or the Tac subunit of the human interleukin-2 receptor (IL-2R) used as a surface protein. Cells expressing either VSV-G or IL-2R were subjected to MACS using the specific antibody-coated magnetic beads. The sorted cells were shown to express predominantly the surface antigen and the CAT. The sensitivity and selectivity of the MACS procedure were further examined by selection of a small subpopulation of cells expressing an endogenous surface protein, the 170-kDa P-glycoprotein which confers the multidrug resistance (MDR1). The low level multidrug-resistant cells could be sorted from mixtures containing drug-sensitive (MDR1-negative) KB-3-1 or bone marrow cells with an efficiency of greater than 70%. The high sensitivity of the method was utilized to screen MDR1-positive cells present in human lymphomas from patients prior to chemotherapy. The percentage of cells isolated by MACS varied from 1 to 20% from the patient samples. The presence of MDR1 expression in these sorted cells was confirmed by the polymerase chain reaction.; In another study, the MACS procedure was used to select monkey kidney (CV-1) cell populations in which the expression of CAT, either transiently or stably, was suppressed by transfection of antisense oligonucleotides in vivo. The results showed that the antisense oligomers were able to inhibit CAT expression in either of the two cases in the sorted cells. The MACS procedure was also used for the development of a novel cell fusion technology applicable to any two cell types. Finally, the MACS method using immunocolloidal ('ferrofluids') system was used to isolate progenitor cells from liver using a surface antigen OV1 unique to these cells.