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EDITORIAL: SPECTROSCOPIC IMAGING
Citation:
LERNER, J., S. LOCKETT, AND R. M. ZUCKER. EDITORIAL: SPECTROSCOPIC IMAGING. CYTOMETRY. Alan R Liss, Inc, New York, NY, 69A(8):711, (2006).
Impact/Purpose:
This is an editorial
Description:
A foremost goal in biology is understanding the molecular basis of single cell behavior, as well as cell interactions that result in functioning tissues. Accomplishing this goal requires quantitative analysis of multiple, specific macromolecules (e.g. proteins, ligands and enzymes) in the spatially- organized environments of intact cells and tissues. This is achieved in practice by fluorescence labeling of the molecular species of interest; using optical microscopy to localize the labeled molecules, and applying computational image analysis for quantifying the molecules in cells or sub-cellular compartments. These approaches have been evolving over the past 30 years, and there are now a wide range of techniques for analyzing the location, dynamics and interactions of molecules in cells. One of these techniques is spectroscopic imaging, which simultaneously quantifies multiple (usually up to five) fluorescence labels with different spectral emission properties at every point in the image. For most people spectroscopic imaging was launched in 1972 with the first Landsat remote Earth sensing satellite. The goal then, as now, was to capture the spectral features of a variety of objects or conditions, such as pollution, minerals and vegetation that could be classified as "spectral objects." The spectral objects are then represented in the images using different pseudo-colors. For example, we have all seen in newspapers and on television how the amount of rainfall, temperature, or pressure can be presented in different colors overlaid on a map of the country. Following the great success of spectroscopic imaging in remote Earth sensing, it was applied to other scientific disciplines, including the field of fluorescence microscopy about 15 years ago. However, spectroscopic imaging only really took off in this field over the last five years when spectral imaging detectors and "un-mixing" algorithms for separating the overlapping spectra of fluorescence labels became commercially available on confocal microscopes. This special issue of Cytometry Part A explains, reviews and reports original research about techniques and applications of spectroscopic imaging mainly in the field of fluorescence imaging of cells. Thus, papers in the issue cover the principals of the optical hardware for collecting emission spectra, computational techniques for un-mixing the spectra of different fluorescence labels, new labels with beneficial spectral properties and methods for calibrating spectroscopic imaging systems. Other papers in the issue report on Raman spectral imaging methods for detecting unlabeled chemical species, as well as papers presenting applications from other scientific disciplines that indicate new possibilities for cell imaging. It is clear that there are still many challenges that need to be resolved before spectroscopic imaging will be a routine tool in every laboratory. For example, there is still considerable inaccuracy in the quantification of specific molecular species when fluorescence spectra significantly overlap or when unknown fluorescence signals such as from autofluorescence are co-mingled with the specific signals of interest. Hence, an outstanding issue is the need for protocols for calibrating and validating the instrumentation used to generate spectral data. Spectroscopic imaging is exciting and dynamic. We hope this issue provides a solid introduction to the field and stimulates technical advances and cytometric applications that augment our understanding of molecular mechanisms driving cell behavior.