Imaging spectrometer fundamentals for researchers in the biosciences--a tutorial.
Over the last 2 years there has been a dramatic increase in the number of bioscience laboratories using wavelength dispersive spectroscopy to study in vivo, in situ fluorescence. Transforming spectral information into an image provides a graphic means of mapping localized ionic, molecular, and protein-protein interactions. Spectroscopy also enables fluorophores with overlapping spectral features to be delineation. In this study, we provide the tools that a researcher needs to put into perspective instrumental contributions to a reported spectrum in order to gain greater understanding of the natural emission of the sample. We also show how to deduce the basic capabilities of a spectral confocal system. Finally, we show how to determine the true spectral bandwidth of an object, the illuminated area of a laser-excited object, and what is needed to optimize light throughput.
Lerner
Jeremy M
JM
LightForm Inc, Hillsborough, NJ 08844, USA. jlerner@lightforminc.com
D000818:Animals; D002138:Calibration; D006801:Humans; D019044:Image Cytometry / Q000295:instrumentation / Q000379:methods*; D008433:Mathematics; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods; D055095:Optics and Photonics; D013050:Spectrometry, Fluorescence / Q000295:instrumentation / Q000379:methods*