Confocal mosaicing microscopy in Mohs skin excisions: feasibility of rapid surgical pathology

Mosaicing of confocal images enables observation of

nuclear morphology in large areas of tissue. An application of interest

is rapid detection of basal cell carcinomas BCCs in skin excisions

during Mohs surgery. A mosaic is currently created in less than 9 min,

whereas preparing frozen histology requires 20 to 45 min for an excision.

In reflectance mosaics, using acetic acid as a contrast agent to

brighten nuclei, large and densely nucleated BCC tumors were detectable

in fields of view of 1212 mm which is equivalent to a

2-magnified view as required by Mohs surgeons. However, small

and sparsely nucleated tumors remained undetectable. Their diminutive

size within the large field of view resulted in weak light backscatter

and contrast relative to the bright surrounding normal dermis. In

fluorescence, a nuclear-specific contrast agent may be used and light

emission collected specifically from nuclei but almost none from the

dermis. Acridine orange of concentration 1 mM stains nuclei in 20 s

with high specificity and strongly enhances nuclear-to-dermis contrast

of BCCs. Comparison of fluorescence mosaics to histology shows that

both large and small tumors are detectable. The results demonstrate

the feasibility of confocal mosaicing microscopy toward rapid surgical

pathology to potentially expedite and guide surgery.