Protein conformation and molecular order probed by second-harmonic-generation microscopy.
Second-harmonic-generation (SHG) microscopy has emerged as a powerful tool to image unstained living tissues and probe their molecular and supramolecular organization. In this article, we review the physical basis of SHG, highlighting how coherent summation of second-harmonic response leads to the sensitivity of polarized SHG to the three-dimensional distribution of emitters within the focal volume. Based on the physical description of the process, we examine experimental applications for probing the molecular organization within a tissue and its alterations in response to different biomedically relevant conditions. We also describe the approach for obtaining information on molecular conformation based on SHG polarization anisotropy measurements and its application to the study of myosin conformation in different physiological states of muscle. The capability of coupling the advantages of nonlinear microscopy (micrometer-scale resolution in deep tissue) with tools for probing molecular structure in vivo renders SHG microscopy an extremely powerful tool for the advancement of biomedical optics, with particular regard to novel technologies for molecular diagnostic in vivo.
University of Florence, Department of Evolutionary Biology Leo Pardi, Florence, Italy.
Francesco
F
Vanzi
Leonardo
L
Sacconi
Riccardo
R
Cicchi
Francesco S
FS
Pavone
D000818:Animals; D016880:Anisotropy; D003094:Collagen / Q000737:chemistry; D003315:Cornea / Q000502:physiology; D006801:Humans; D008853:Microscopy / Q000379:methods*; D008859:Microscopy, Polarization / Q000379:methods; D015233:Models, Statistical; D008968:Molecular Conformation; D015394:Molecular Structure; D009132:Muscles / Q000166:cytology; D009218:Myosins / Q000737:chemistry; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry; D012542:Scattering, Radiation; D013552:Swine
D011506:Proteins; D003094:Collagen; D009218:Myosins