Second harmonic generation (SHG) has been developed in our laboratories as
a high-resolution nonlinear optical imaging microscopy for cellular membran
es and intact tissues. SHG shares many of the advantageous features for mic
roscopy of another more established nonlinear optical technique: two-photon
excited fluorescence (TPEF). Both are capable of optical sectioning to pro
duce three-dimensional images of thick specimens and both result in less ph
otodamage to living tissue than confocal microscopy. SHG is complementary t
o TPEF in that it uses a different contrast mechanism and is most easily de
tected in the transmitted light optical path. It can be used to image membr
ane probes with high membrane specificity and displays extraordinary sensit
ivity in reporting membrane potential; it also has the ability to image hig
hly ordered structural proteins without any exogenous labels. (C) 2001 Soci
ety of Photo-Optical Instrumentation Engineers.