Characterization of phytoplankton community composition is critical to understanding the ecology and biogeochemistry of the oceans. One approach to taxonomic characterizationtakes advantage of differing pigmentation between algal taxa and thus differences in fluorescence excitation spectra. Analyses of bulk water samples, however, may be confounded by interference from chromophoric dissolved organic matter or suspended particulate matter. Here, we describe an instrument that uses a laser trap based on a Nikon TE2000-U microscope to position individual phytoplankton cells for confocal fluorescence excitation spectroscopy, thus avoiding interference from the surrounding medium. Quantitative measurements of optical power give data in the form of photonsemitted per photon of exposure for an individual phytoplankton cell. Residence times for individual phytoplankton in the instrument can be as long as several minutes with no substantial change in their fluorescence excitation spectra. The laser trap was found togenerate two-photon fluorescence from the organisms so a modification was made to release the trap momentarily during data acquisition. Typical signal levels for an individual cell are in the range of 106 photons/s of fluorescence using a monochromated 75 W Xe arc lamp excitation source with a 2% transmission neutral density filter.
Thursday, February 11, 2010
Construction, figures of merit, and testing of a single-cell fluorescence excitation spectroscopy system
Laura S. Hill, Tammi L. Richardson, Luisa T. M. Profeta, Timothy J. Shaw, Christopher J. Hintz, Benjamin S. Twining, Evelyn Lawrenz, and Michael L. Myrick
Characterization of phytoplankton community composition is critical to understanding the ecology and biogeochemistry of the oceans. One approach to taxonomic characterizationtakes advantage of differing pigmentation between algal taxa and thus differences in fluorescence excitation spectra. Analyses of bulk water samples, however, may be confounded by interference from chromophoric dissolved organic matter or suspended particulate matter. Here, we describe an instrument that uses a laser trap based on a Nikon TE2000-U microscope to position individual phytoplankton cells for confocal fluorescence excitation spectroscopy, thus avoiding interference from the surrounding medium. Quantitative measurements of optical power give data in the form of photonsemitted per photon of exposure for an individual phytoplankton cell. Residence times for individual phytoplankton in the instrument can be as long as several minutes with no substantial change in their fluorescence excitation spectra. The laser trap was found togenerate two-photon fluorescence from the organisms so a modification was made to release the trap momentarily during data acquisition. Typical signal levels for an individual cell are in the range of 106 photons/s of fluorescence using a monochromated 75 W Xe arc lamp excitation source with a 2% transmission neutral density filter.
Characterization of phytoplankton community composition is critical to understanding the ecology and biogeochemistry of the oceans. One approach to taxonomic characterizationtakes advantage of differing pigmentation between algal taxa and thus differences in fluorescence excitation spectra. Analyses of bulk water samples, however, may be confounded by interference from chromophoric dissolved organic matter or suspended particulate matter. Here, we describe an instrument that uses a laser trap based on a Nikon TE2000-U microscope to position individual phytoplankton cells for confocal fluorescence excitation spectroscopy, thus avoiding interference from the surrounding medium. Quantitative measurements of optical power give data in the form of photonsemitted per photon of exposure for an individual phytoplankton cell. Residence times for individual phytoplankton in the instrument can be as long as several minutes with no substantial change in their fluorescence excitation spectra. The laser trap was found togenerate two-photon fluorescence from the organisms so a modification was made to release the trap momentarily during data acquisition. Typical signal levels for an individual cell are in the range of 106 photons/s of fluorescence using a monochromated 75 W Xe arc lamp excitation source with a 2% transmission neutral density filter.
Subscribe to:
Post Comments (Atom)
No comments:
Post a Comment