A study was performed to evaluate in vitro the sensitivity, specificity and variability of a new immunomagnetic
microbead isolation technique which provides subsequent immunological staining of captured
carcinoma cells. In a mixture of peripheral blood mononuclear cells (PBMCs) and human
carcinoma cells the epithelial
cancer cells were isolated with the Dynal((R)) RAM
IgG1 CELLection Kit using Dynabeads
M-280 coated with a rat
monoclonal antibody (Mab) against mouse
IgG1. The rat Mab was biotinylated and attached to Dynabeads via
streptavidin and
a DNA linker. The anti-epithelial monoclonal mouse antibody Ber-EP4 was used as the primary capture antibody. In order to permit phenotyping of the isolated
carcinoma cells the magnetic beads were removed from the
carcinoma cells by DN'ase digestion of the
DNA linker between the magnetic bead and the secondary antibody. In an ex vivo model system an average recovery of approximately 60% of a human colon
carcinoma cell line HCC-2998 seeded in 5.10(6) PBMCs was obtained, and the recovered cells could subsequently be immunologically stained for the
surface antigen CD87 (
urokinase plasminogen activator receptor). No positive stained cells were found in control experiments with PBMCs without
carcinoma cells. Despite a relatively low recovery, the described method will be valuable for the detection of
carcinoma cells in cytospin preparations with subsequent phenotyping of the cells for expression of
surface antigens. Depending on the chosen
antibodies, the method may be useful for the isolation and characterisation of other cell types in various cell
suspensions.