Despite many successes, atomic force microscopy (AFM) of biological sp
ecimens at room temperature is still severely limited by at least two
factors: the softness and the thermal motion of flexible multi-domain/
subunit molecules. Both problems can be overcome by imaging biological
structures at cryogenic temperatures. Even though the instrumentation
is considerably more complex and earlier attempts were largely unsucc
essful, cryo-AFM has recently been demonstrated on a number of biologi
cal specimens, using an AFM operated in liquid nitrogen vapor under am
bient pressure. In this brief review, both the method of instrumentati
on and the latest biological applications are discussed. Not only has
the cryo-AFM attained high resolution on those specimens that could no
t be well imaged at room temperature, but it has also produced potenti
ally important information on several specimens. These results firmly
establish the cryo-AFM as a useful and versatile structural probe in b
iology with its own unique capabilities.