MOLECULAR analysis of complex biological structures and processes incr
easingly requires sensitive methods far protein sequencing. Electrospr
ay mass spectrometry(1) has been applied to the high-sensitivity seque
ncing of short peptides(2), but technical difficulties have prevented
similar success with gel-isolated proteins. Here we report a simple an
d robust technique for the sequencing of proteins isolated by polyacry
lamide gel electrophoresis, using nano-electrospray(3,4) tandem mass s
pectrometry(5,6). As little as 5 ng protein starting material on Cooma
ssie- or silver-stained gels can be sequenced. Multiple-sequence stret
ches of up to 16 amino acids are obtained, which identify the protein
unambiguously if already present in databases or provide information t
o clone the corresponding gene. We have applied this method to the seq
uencing and cloning of a protein which inhibits the proliferation of c
apillary endothelial cells in vitro and thus may have potential antian
giogenic effects on solid tumours.