Figure 4

Lectin-based cell-selection strategies. Two lectin-based strategies can separate cells with glycosylation abnormalities from predominantly wild-type populations. (a) The first strategy uses a bifunctional toxic lectin that binds to a carbohydrate ligand expressed on wild-type cells thereby causing cell death. Reduced (Mutant cell 1) or altered (Mutant cell 2) expression of the target epitope can abrogate lectin binding allowing cell recovery. A drawback of this strategy is that over-expression phenotypes (Mutant cell 3) are targeted for death, preventing their isolation; in addition, intermediate levels of lectin binding (Mutant cell 1) may or may not cause cell death. (b)A superior selection strategy uses fluorescently labeled lectins that allow separation of cell types by flow cytometry (FACS). In this case, sorting conditions are established with wild-type cells and these subsequently allow separation of cells with low, no, or even excess, lectin binding by using low, negative, or high FACS parameters. As a result, each of the mutant cell populations can be isolated with a level of precision not readily attained with the toxic lectin approach.