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Histopathology, whether visualizing microstructure or selectively labeling molecules with special stains, has a long history
of development and maturation, and has been instrumental in biological or clinical laboratories for basic research and
in hospitals for disease diagnosis/screening. To avoid the artifacts associated with sample freezing, fixation, and staining
(labeling), we develop a nonlinear penta-model spectro-microscopy to critically complement or potentially eliminate stained
histology and histopathology. This technique requires a shift of focus from manipulating matter (the biological sample)
to instead manipulating the optical waves (via customized excitation and signal detection) in order to generate molecular
contrast. Instead of using the exogenous labeling agents in conventional optical microscopy to “physically” (invasively) label
the chemical substances of interest; we initiate the use of rapidly switchable light excitation/detection channels to virtually
“label” these chemical substances. Thus, the biological sample can be visualized in its physiologically authentic condition
without sacrificing either molecular specificity or high spatial resolution. This technique is implemented in a programmable
microscope requiring minimum optical realignment, so that a biologist or pathologist with no laser training can in the future
selectively display a specific endogenous molecule (or molecular structure) on the computer screen by programming the
excitation, or instantly compare the displayed molecule distribution with a different one by reprogramming the excitation.
Using breast cancer as a prototypical application, we have imaged the well-known events in tumor microenvironment,
including angiogenesis, lymphangiogenesis, extracellular matrix remodeling, non-native cell recruitment, extracellular vesicle
up-production and switched metabolism toward biosynthesis. This allows us to identify early and quantifiable biomarkers in
breast cancer development.