Visualization of leukocyte transendothelial and interstitial migration using reflected light oblique transillumination in intravital video microscopy

Dynamic visualization of the intravascular events leading to the
extravasation of leukocytes into tissues by intravital microscopy
has significantly expanded our understanding of the underlying
molecular processes. In contrast, the detailed observation of
leukocyte transendothelial and interstitial migration in vivo has
been hampered by the poor image contrast of cells within turbid
media that is obtainable by conventional brightfield microscopy.
Here we present a microscopic method, termed reflected light
oblique transillumination microscopy, that makes use of the optical
interference phenomena generated by oblique transillumination to
visualize subtle gradients of refractive indices within tissues for
enhanced image contrast. Using the mouse cremaster muscle, we
demonstrate that this technique makes possible the reliable
quantification of extravasated leukocytes as well as the
characterization of morphological phenomena of leukocyte
transendothelial and interstitial migration.