ABSTRACT
Spatial resolution, within a microscopic field of view, is the primary achievement of the optically efficient, modem infrared microspectrometer. The chemistry of single cells in biological specimens can be studied in situ using an infrared spec troscopic probe of cellular dimensions. Infrared microspectroscopy combines the fields of infrared spectroscopy, microscopy, and computer science. The result enables a comparison of microspectroscopic chemical information to histological structures. In light microscopy, image contrast is produced by the application of stains or fluorescent materials. Electronic microspectroscopy produces spectra of individual pixels or select wavelength images. In vibrational microspectroscopy (infrared or Raman), the use of chemical reagents or stains is not necessary. In mapping procedures with microbeam molecular spectroscopy, the contrast in images produced is from intrinsic infrared absorption bands. From multiple probing, functional-group maps can be established from baseline-corrected ab sorbance values (peak height or area). In this way, the spectral and chemical integrities of the tissue being studied are not compromised by the elimination of homogenization and chemical modifications via staining. Fourier transform infrared (FT-IR) microspectroscopy does more than perform microanalysis on small samples; it allows spatially resolved localized chemical analysis in situ from small portions of the microscopic field, thereby relating localized chemical analysis to the morphology (histology) of the specimen. The presence of certain
organic groups is established or excluded by looking for their particular intrinsic absorption bands. Displaying the spectrum in absorbance allows the magnitude of the absorbance reading to be related to the relative concentration of those particular organic groups.
