Microcontact printing of axon guidance molecules for generation of graded patterns.
Microcontact printing (microCP) of proteins has been successfully used for patterning surfaces in various contexts. Here we describe a simple 'lift-off' method to print precise patterns of axon guidance molecules, which are used as substrate for growing chick retinal ganglion cell (RGC) axons. Briefly, the etched pattern of a silicon master is transferred to a protein-coated silicone cuboid (made from polydimethylsiloxane, PDMS), which is then used as a stamp on a glass coverslip. RGC explants are placed adjacent to the pattern and cultured overnight. Fluorescent labeling of the printed proteins allows the quantitative analysis of the interaction of axons and growth cones with single protein dots and of the overall outgrowth and guidance rate in variously designed patterns. Patterned substrates can be produced in 3-4 h and are stable for up to one week at 4 degrees C; the entire protocol can be completed in 3 d.
von Philipsborn
Anne C
AC
Max-Planck-Institut für Entwicklungsbiologie, Spemannstrasse 35, 72076 Tübingen, Germany.
Lang
Susanne
S
Bernard
André
A
Loeschinger
Jürgen
J
David
Christian
C
Lehnert
Dirk
D
Bastmeyer
Martin
M
Bonhoeffer
Friedrich
F
D000818:Animals; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D002645:Chickens; D003952:Diagnostic Imaging; D004129:Dimethylpolysiloxanes; D005456:Fluorescent Dyes; D020439:Growth Cones / Q000737:chemistry* / Q000648:ultrastructure*; D009419:Nerve Tissue Proteins / Q000648:ultrastructure*; D012165:Retinal Ganglion Cells / Q000166:cytology*
D004129:Dimethylpolysiloxanes; D005456:Fluorescent Dyes; D009419:Nerve Tissue Proteins