Correlative Synchrotron Fourier Transform Infrared Spectroscopy and Single Molecule Super Resolution Microscopy for the Detection of Composition and Ultrastructure Alterations in Single Cells

Single molecule localization microscopy (SMLM) and synchrotron Fourier transform infrared (S-FTIR) spectroscopy are two techniques capable of elucidating unique and valuable biological detail. SMLM provides images of the structures and distributions of targeted biomolecules at spatial resolutions up to an order of magnitude better than the diffraction limit, whereas IR spectroscopy objectively measures the holistic biochemistry of an entire sample, thereby revealing any variations in overall composition. Both tools are currently applied extensively to detect cellular response to disease, chemical treatment, and environmental change. Here, these two techniques have been applied correlatively at the single cell level to probe the biochemistry of common fixation methods and have detected various fixation-induced losses of biomolecular composition and cellular ultrastructure. Furthermore, by extensive honing and optimizing of fixation protocols, many fixation artifacts previously considered pervasive and regularly identified using IR spectroscopy and fluorescence techniques have been avoided. Both paraformaldehyde and two-step glutaraldehyde fixation were identified as best preserving biochemistry for both SMLM and IR studies while other glutaraldehyde and methanol fixation protocols were demonstrated to cause significant biochemical changes and higher variability between samples. Moreover, the potential complementarity of the two techniques was strikingly demonstrated in the correlated detection of biochemical changes as well as in the detection of fixation-induced damage that was only revealed by one of the two techniques

Assessment of River Herring and Striped Bass in the Connecticut River: Abundance, Population Structure, and Predator/Prey Interactions

Populations of anadromous alewife Alosa pseudoharengus and blueback herring A. aestivalis, collectively referred to as river herring, have declined in the Connecticut River. An explanatory hypothesis for these declines is that predation pressures have increased as a result of recent increases in abundance of sympatric striped bass Morone saxatilis. We sampled river herring and striped bass from the stretch of the Connecticut River between Wethersfield, CT and Holyoke, MA during the vernal migration seasons of 2005-2008. The objectives of the sampling program were to assess abundance, temporal/spatial distribution, and population structure of both river herring and striped bass, as well as striped bass food habits. Blueback herring population structure has changed over recent decades. Contemporary runs feature younger, smaller fish that are less likely to complete multiple spawning runs over their lifetime. These temporal shifts are indicative of elevated mortality rates operating on older, larger herring. Striped bass predation is a significant source of mortality for adult blueback herring in the Connecticut River. River herring comprise a significant portion of striped bass diets in the Connecticut River during May-June, and striped bass congregate in locations where they are successful in capturing herring. The estimated seasonal consumption of blueback herring by striped bass in our study stretch is comparable to the numbers of herring passed annually at the Holyoke fish lift prior to the onset of recent declines. Future studies will incoporate estimates of predation mortality described here into structured population models that can be used to hindcast the impact of striped bass predation on river herring run size in recent decades, and examine the potential for amelioration of river herring mortality via changes to management of striped bass fisheries

Band assignments for the liver tissue spectra.

<p>Band assignments for the liver tissue spectra.</p

Second derivative spectra from Fig. 3 for liver tissue: hydrated tissue ATR spectrum (blue), formalin fixed transmission spectrum (black), desiccator dried transmission spectrum (pink) and ethanol dehydrated transmission spectrum (green).

<p>Second derivative spectra from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0116491#pone.0116491.g003" target="_blank">Fig. 3</a> for liver tissue: hydrated tissue ATR spectrum (blue), formalin fixed transmission spectrum (black), desiccator dried transmission spectrum (pink) and ethanol dehydrated transmission spectrum (green).</p