Table 1.

Comparative features between Hematoxylin-Eosin (H&E) and Multiphoton Microscopy (MPM) for evaluation of human bladder biopsies.

	Diagnostic features identifiable on both H&E and Multiphoton microscopy (MPM)	Multiphoton microscopy (MPM) limitations
Benign, flat intraurothelial lesions	Multilayered urothelium with superficial umbrella cells Maintained cell polarity Underlying lamina propria with blood vessels, von Brunn nests	Lack of intranuclear details
Carcinoma in situ (CIS)	Flat lesion Marked pleomorphism High nuclear:cytoplasmic (N:C) ratio Loss of polarity	Lack of intranuclear detail may impair ability to distinguish CIS from urothelial dysplasia or reactive urothelium
Low grade papillary urothelial carcinoma	Papillary fronds with thin fibrovascular cores Overall orderly arrangement of urothelial cells with mild pleomorphism Relatively low Nuclear:Cytoplasmic (N:C) ratio	Difficult to differentiate between papilloma, papillary urothelial neoplasms of low malignant potential (PUNLMP) and low grade papillary urothelial carcinoma
High grade papillary urothelial carcinoma	Papillary fronds and occasional sheets of tumor cells Moderate to marked nuclear and cellular pleomorphism High Nuclear:Cytoplasmic (N:C) ratio Loss of polarity	Lack of intranuclear detail may impair ability to distinguish high grade from low grade lesions
Invasive urothelial carcinoma	Irregular nests of tumor cells, or individual tumor cells, within lamina propria (pT1) or muscularis propria (pT2)	Invasion reliably identified only if extensive; microinvasion not easily identified Difficult to assess muscularis propria presence, including involvement by tumor, due to penetration depth limitation in current Multiphoton microscopy (MPM) imaging instrumentation