Specific Selection of Mitotically Active Intestinal Cells by Concanavalin A-derivatized Fibers

INTRODUCTION of [3H]thymidine) may be isolated from the nonincorporating villus cells . After obtaining cells, the Concanavalin A (Con A), a lectin isolated from separate villus and crypt cell fractions were washed the jack bean, is known to preferentially agglutiseveral times in phosphate-buffered 0 .154 N NaCl nate certain cells (1) . Preferential agglutination solution (PBS) and subsequently filtered through silk by Con A has been demonstrated for virally and glass wool to yield a suspension of single cells . transformed tissue culture cells, chick embryo Cell concentrations were adjusted to 5 X 10 6 cells/ retinal cells (2), and human fetal intestinal cells ml . (3) . Although the mechanism of agglutination is Con A was covalently bound to nylon fibers using still not clear, agglutination by Con A appears a carbodiimide condensing agent (1-cyclohexyl-3]2to require an interaction of the lectin with the inorpholinoethyl]-carbodiimide-metho p-toluenesulfocarbohydrate portion of cell surface membrane nate) according to the method of Edelman et al . (6) . The cells were put into small plastic petri dishes glycoproteins (4) . This interaction (or binding) (Falcon Plastics, Division of B-D Laboratories, Inc . to cell surface membrane glycoproteins which Los Angeles, Calif.) (diam 33 mm) and incubated leads to agglutination appears to be relatively with the Con A-derivatized fibers at room temperaspecific for undifferentiated (or dedifferentiated) ture for 10 min with constant shaking (120 oscillamitotically active cells. The following experitions/min) . After 10 min the fibers were removed and ments support this relative specificity by demonwashed for 2 min in petri dishes containing PBS . strating that Con A preferentially selected a popuThen the fibers were placed in petri dishes containing lation of rat intestinal epithelial cells which appear a-methylinannoside (100 mM), a potent competitive to be undifferentiated crypt cells . inhibitor of Con A binding . Incubation with the inhibitor caused release of better than 90% of the