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Fluorescence measurements for evaluating the application of multivariate analysis techniques to optically thick environments

Reichardt, Thomas A.; Schmitt, Randal L.; Sickafoose, Shane S.; Jones, Howland D.; Timlin, Jerilyn A.

Laser-induced fluorescence measurements of cuvette-contained laser dye mixtures are made for evaluation of multivariate analysis techniques to optically thick environments. Nine mixtures of Coumarin 500 and Rhodamine 610 are analyzed, as well as the pure dyes. For each sample, the cuvette is positioned on a two-axis translation stage to allow the interrogation at different spatial locations, allowing the examination of both primary (absorption of the laser light) and secondary (absorption of the fluorescence) inner filter effects. In addition to these expected inner filter effects, we find evidence that a portion of the absorbed fluorescence is re-emitted. A total of 688 spectra are acquired for the evaluation of multivariate analysis approaches to account for nonlinear effects.

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From benchtop to raceway : spectroscopic signatures of dynamic biological processes in algal communities

Timlin, Jerilyn A.; Garcia, Omar F.; Aragon, Michelle L.; Powell, Amy J.; Jones, Howland D.; Reichardt, Thomas A.; Ricken, James B.; Trahan, Christine A.; Ruffing, Anne R.; Collins, Aaron M.; Dwyer, Brian P.

The search is on for new renewable energy and algal-derived biofuel is a critical piece in the multi-faceted renewable energy puzzle. It has 30x more oil than any terrestrial oilseed crop, ideal composition for biodiesel, no competition with food crops, can be grown in waste water, and is cleaner than petroleum based fuels. This project discusses these three goals: (1) Conduct fundamental research into the effects that dynamic biotic and abiotic stressors have on algal growth and lipid production - Genomics/Transcriptomics, Bioanalytical spectroscopy/Chemical imaging; (2) Discover spectral signatures for algal health at the benchtop and greenhouse scale - Remote sensing, Bioanalytical spectroscopy; and (3) Develop computational model for algal growth and productivity at the raceway scale - Computational modeling.

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Use of ceregenins to create novel biofouling resistant water water-treatment membranes

Altman, Susan J.; Jones, Howland D.; Kirk, Matthew F.

Scoping studies have demonstrated that ceragenins, when linked to water-treatment membranes have the potential to create biofouling resistant water-treatment membranes. Ceragenins are synthetically produced molecules that mimic antimicrobial peptides. Evidence includes measurements of CSA-13 prohibiting the growth of and killing planktonic Pseudomonas fluorescens. In addition, imaging of biofilms that were in contact of a ceragenin showed more dead cells relative to live cells than in a biofilm that had not been treated with a ceragenin. This work has demonstrated that ceragenins can be attached to polyamide reverse osmosis (RO) membranes, though work needs to improve the uniformity of the attachment. Finally, methods have been developed to use hyperspectral imaging with multivariate curve resolution to view ceragenins attached to the RO membrane. Future work will be conducted to better attach the ceragenin to the RO membranes and more completely test the biocidal effectiveness of the ceragenins on the membranes.

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Optimizing algal cultivation & productivity : an innovative, multidiscipline, and multiscale approach

Timlin, Jerilyn A.; Jones, Howland D.; Ricken, James B.; Murton, Jaclyn K.; Dwyer, Brian P.; Ruffing, Anne R.; Powell, Amy J.; Reichardt, Thomas A.

Progress in algal biofuels has been limited by significant knowledge gaps in algal biology, particularly as they relate to scale-up. To address this we are investigating how culture composition dynamics (light as well as biotic and abiotic stressors) describe key biochemical indicators of algal health: growth rate, photosynthetic electron transport, and lipid production. Our approach combines traditional algal physiology with genomics, bioanalytical spectroscopy, chemical imaging, remote sensing, and computational modeling to provide an improved fundamental understanding of algal cell biology across multiple cultures scales. This work spans investigations from the single-cell level to ensemble measurements of algal cell cultures at the laboratory benchtop to large greenhouse scale (175 gal). We will discuss the advantages of this novel, multidisciplinary strategy and emphasize the importance of developing an integrated toolkit to provide sensitive, selective methods for detecting early fluctuations in algal health, productivity, and population diversity. Progress in several areas will be summarized including identification of spectroscopic signatures for algal culture composition, stress level, and lipid production enabled by non-invasive spectroscopic monitoring of the photosynthetic and photoprotective pigments at the single-cell and bulk-culture scales. Early experiments compare and contrast the well-studied green algae chlamydomonas with two potential production strains of microalgae, nannochloropsis and dunnaliella, under optimal and stressed conditions. This integrated approach has the potential for broad impact on algal biofuels and bioenergy and several of these opportunities will be discussed.

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Analysis of micromixers and biocidal coatings on water-treatment membranes to minimize biofouling

Altman, Susan J.; Clem, Paul G.; Cook, Adam W.; Hart, William E.; Ho, Clifford K.; Jones, Howland D.; Sun, Amy C.; Webb, Stephen W.

Biofouling, the unwanted growth of biofilms on a surface, of water-treatment membranes negatively impacts in desalination and water treatment. With biofouling there is a decrease in permeate production, degradation of permeate water quality, and an increase in energy expenditure due to increased cross-flow pressure needed. To date, a universal successful and cost-effect method for controlling biofouling has not been implemented. The overall goal of the work described in this report was to use high-performance computing to direct polymer, material, and biological research to create the next generation of water-treatment membranes. Both physical (micromixers - UV-curable epoxy traces printed on the surface of a water-treatment membrane that promote chaotic mixing) and chemical (quaternary ammonium groups) modifications of the membranes for the purpose of increasing resistance to biofouling were evaluated. Creation of low-cost, efficient water-treatment membranes helps assure the availability of fresh water for human use, a growing need in both the U. S. and the world.

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Accurate measurement of cellular autofluorescence is critical for imaging of host-pathogen interactions

Progress in Biomedical Optics and Imaging - Proceedings of SPIE

Timlin, Jerilyn A.; Noek, Rachel M.; Kaiser, Julia N.; Sinclair, Michael B.; Jones, Howland D.; Davis, Ryan W.; Lane, Todd L.

Cellular autofluorescence, though ubiquitous when imaging cells and tissues, is often assumed to be small in comparison to the signal of interest. Uniform estimates of autofluorescence intensity obtained from separate control specimens are commonly employed to correct for autofluorescence. While these may be sufficient for high signal-to-background applications, improvements in detector and probe technologies and introduction of spectral imaging microscopes have increased the sensitivity of fluorescence imaging methods, exposing the possibility of effectively probing the low signal-to-background regime. With spectral imaging, reliable monitoring of signals near or even below the noise levels of the microscope is possible if autofluorescence and background signals can be accurately compensated for. We demonstrate the importance of accurate autofluorescence determination and utility of spectral imaging and multivariate analysis methods using a case study focusing on fluorescence confocal spectral imaging of host-pathogen interactions. In this application fluorescent proteins are produced when bacteria invade host cells. Unfortunately the analyte signal is spectrally overlapped and typically weaker than the cellular autofluorescence. In addition to discussing the advantages of spectral imaging for following pathogen invasion, we present the spectral properties of mouse macrophage autofluorescence. The imaging and analysis methods developed are widely applicable to cell and tissue imaging. © 2008 Copyright SPIE - The International Society for Optical Engineering.

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Use of ceragenins to create novel biofouling resistant water-treatment membranes

Altman, Susan J.; Jones, Howland D.; Fellows, Benjamin D.

Scoping studies have demonstrated that ceragenins, when linked to water-treatment membranes have the potential to create biofouling resistant water-treatment membranes. Ceragenins are synthetically produced molecules that mimic antimicrobial peptides. Evidence includes measurements of CSA-13 prohibiting the growth of and killing planktonic Pseudomonas fluorescens. In addition, imaging of biofilms that were in contact of a ceragenin showed more dead cells relative to live cells than in a biofilm that had not been treated with a ceragenin. This work has demonstrated that ceragenins can be attached to polyamide reverse osmosis (RO) membranes, though work needs to improve the uniformity of the attachment. Finally, methods have been developed to use hyperspectral imaging with multivariate curve resolution to view ceragenins attached to the RO membrane. Future work will be conducted to better attach the ceragenin to the RO membranes and more completely test the biocidal effectiveness of the ceragenins on the membranes.

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Weighting hyperspectral image data for improved multivariate curve resolution results

Journal of Chemometrics

Jones, Howland D.; Haaland, David M.; Sinclair, Michael B.; Melgaard, David K.; Van Benthem, Mark V.; Pedroso, M.C.

The combination of hyperspectral confocal fluorescence microscopy and multivariate curve resolution (MCR) provides an ideal system for improved quantitative imaging when multiple fluorophores are present. However, the presence of multiple noise sources limits the ability of MCR to accurately extract pure-component spectra when there is high spectral and/or spatial overlap between multiple fluorophores. Previously, MCR results were improved by weighting the spectral images for Poisson-distributed noise, but additional noise sources are often present. We have identified and quantified all the major noise sources in hyperspectral fluorescence images. Two primary noise sources were found: Poisson-distributed noise and detector-read noise. We present methods to quantify detector-read noise variance and to empirically determine the electron multiplying CCD (EMCCD) gain factor required to compute the Poisson noise variance. We have found that properly weighting spectral image data to account for both noise sources improved MCR accuracy. In this paper, we demonstrate three weighting schemes applied to a real hyperspectral corn leaf image and to simulated data based upon this same image. MCR applied to both real and simulated hyperspectral images weighted to compensate for the two major noise sources greatly improved the extracted pure emission spectra and their concentrations relative to MCR with either unweighted or Poisson-only weighted data. Thus, properly identifying and accounting for the major noise sources in hyperspectral images can serve to improve the MCR results. These methods are very general and can be applied to the multivariate analysis of spectral images whenever CCD or EMCCD detectors are used. Copyright © 2008 John Wiley & Sons, Ltd.

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Trilinear analysis of images obtained with a hyperspectral imaging confocal microscope

Journal of Chemometrics

Van Benthem, Mark H.; Keenan, Michael R.; Davis, Ryan W.; Liu, Ping; Jones, Howland D.; Haaland, David M.; Sinclair, Michael B.; Brasier, Allan R.

Hyperspectral imaging confocal microscopy (HSI-CM) is a powerful tool for the analysis of cellular processes such as the immune response. HSI-CM is a data rich technique that routinely generates two-way data having a spectral domain and an image or concentration domain. Using a variety of modifications to the instrument or experimental protocols, one can readily produce three-way data with HSI-CM. These data are often amenable to trilinear analysis. For example we have used a time series of 18 images acquired during photobleaching of the fluorophores in an effort to identify fluorescence resonance energy transfer (FRET). The resulting images represent intensity as a function of concentration, wavelength and photodegradation in time, to which we apply our techniques of trilinear decomposition. We have successfully employed trilinear decomposition of photobleaching spectral image data from fixed A549 cells transfected with yellow and green fluorescent proteins (YFP and GFP) as molecular probes of cellular proteins involved in the cellular immune response. While useful in the interpretation biological processes, the size of the data generated with the HSI-CM can be difficult to manage computationally. The 208 x 204 x 512 x 18 elements in the image data require careful processing and efficient analysis algorithms. Accordingly, we have implemented fast algorithms that can quickly perform the trilinear decomposition. In this paper we describe how three-way data are produced and the methods we have used to process them. Specifically, we show that co-adding spectra in a spatial neighborhood is a highly effective method for improving the performance of these algorithms without sacrificing resolution. Copyright © 2008 John Wiley & Sons, Ltd.

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Results 51–75 of 85
Results 51–75 of 85