Spotted substrates for focal presentation of proteins to cells.
Neurites and other cell processes encounter localized deposits of signaling factors as they grow. The difficulty in generating patterned artificial substrates has hindered the analysis of these instructive factors in vitro. Here we report a simple method for presenting cultured cells with small spots of protein on an otherwise uniform substrate. We use a biolistic device called a gene gun to deposit 0.1-5 microm fluorescent dots of pure protein on or beneath a growth-promoting substrate. Using this technique, we demonstrate local presynaptic differentiation of motoneurons in response to dots of a polycation. We also show that biotin-avidin and antibody-antigen interactions can be used to prepare spots from more dilute, more labile, or less abundant proteins. This method should prove useful for analyzing extracellular signaling molecules that act focally on neurons or other cell types.
Kummer
Terrance T
TT
Washington University School of Medicine, St. Louis, MO 63110, USA.
Nishimune
Hiroshi
H
Wong
Rachel O L
RO
Sanes
Joshua R
JR
D000818:Animals; D000937:Antigen-Antibody Reactions; D001360:Avidin / Q000494:pharmacology; D019470:Biolistics / Q000295:instrumentation*; D001710:Biotin / Q000494:pharmacology; D002417:Cattle; D018929:Cell Culture Techniques / Q000295:instrumentation* / Q000379:methods; D002478:Cells, Cultured / Q000187:drug effects; D018397:Equidae; D006041:Goats; D007074:Immunoglobulin G / Q000494:pharmacology; D008567:Membranes, Artificial; D051379:Mice; D008863:Microspheres; D009046:Motor Neurons / Q000166:cytology / Q000187:drug effects*; D016501:Neurites / Q000187:drug effects / Q000648:ultrastructure; D011076:Polycarboxylate Cement; D011107:Polylysine / Q000494:pharmacology; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D013045:Species Specificity; D019809:Streptavidin / Q000494:pharmacology
D007074:Immunoglobulin G; D008567:Membranes, Artificial; D011076:Polycarboxylate Cement; D001360:Avidin; D011107:Polylysine; C005506:polycarbonate; D012710:Serum Albumin, Bovine; D001710:Biotin; D019809:Streptavidin