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The Biosystems Research department is conducting research in the following areas :

More detail for each subject area can be found by exploring the "Researchers" link to the left.

Biodefense

left image is A, right image is B

(a) Late stage of infection of RAW 264.7 macrophages with Francisella novicida Utah 112 expressing fluorescent reporters (b) A microchip used to study cell signaling in immune cells infected with bacteria. Pictures courtesy of Todd Lane (panel a) and Nimisha Srivastava (panel b).

We are one of the leading departments at Sandia involved in basic and applied research in biodefense. Our efforts span basic research in host-pathogen interactions and studying virulence mechanisms of pathogens as well as applied research in developing medical diagnostic and biosensors for biological agents, high-throughput screening for therapeutic targets and proteomic platforms for biomarker discovery and verification.

We lead a multi-disciplinary and multi-departmental grand-challenge project (Microscale Immune Studies laboratory; PDF, 156Kb) to develop novel experimental and computational tools to elucidate the molecular mechanisms of the innate immune response to pathogens such as bacteria and viruses at the single cell level. Our approach utilizes a high-throughput microfluidic platform to provide a controlled environment for manipulating cells and to perform high-resolution hyperspectral confocal imaging and proteomic analysis. Predictive computational modeling is then used to integrate the data to construct a system-level understanding of innate immunity pathways. Our microengineered platform provides the biological research community with a miniaturized, versatile, and modular tool to perform single-cell analysis for many applications including infectious disease research and drug discovery.

In the news

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Biofuels

The development of cheap renewable alternatives to fossil fuel derived transportation fuels would reduce our dependence on imported petroleum. “Energy crops” and agriculture waste are preferred long-term solutions for renewable, cheap and globally available feedstocks for biofuels. Researchers in our group are developing novel technologies and processes for production of biofuels production from cellulosic biomass. Our approach combines computational modeling, genomics, protein engineering and synthetic biology of extremophilic organisms to address the enzymatic breakdown and fermentation of cellulosic biomass. In additon it has been recognized that microalgal biodiesel has the potential of replacing much of the heavy transportation fuel. The development of a method by which algal biodiesel can be produced with high efficiency and low cost would be a disruptive development that would have far-reaching impact. The major bottleneck today is the triacylglyceride production and conversion. We are investigating and developing new techniques for the extraction and conversion of triacylglycerides as well as developing algae with increased triacylglyceride content.

Our researchers are also playing key roles in the recently established Joint Bio-energy Institute (JBEI) funded by Office of Science in the US Department of Energy. For more information on JBEI, please visit http://jbei.lbl.gov

Medical Diagnostics

Using microfluidic chip-based assays, researchers in our group are developing portable devices for early diagnosis of diseases and exposure to pathogens. These devices are geared towards first-responders as well as healthcare professionals in primary care. Only a tiny drop of blood or saliva is required and results can be obtained in less than 10 minutes. The fully automated, self-contained device is controlled by either a laptop computer or a touch screen on the device. Test results digitally indicate the:

  1. presence of disease biomarkers of interest and
  2. relative concentration of said biomarkers in health and disease states.

A prototype weighing approximately 5 lbs. being developed for point-of-care diagnostics

A prototype weighing approximately 5 lbs. being developed for point-of-care diagnostics

The heart of the device is a microfluidic chip that, in a few minutes, performs electrophoretic immunoassays prior to sensitive (~10-12 molar detection limit) laser-induced fluorescence detection requiring only a few microliters of sample. The chip is integrated with miniaturized electronics, optical elements, fluid-handling components, and data acquisition software to provide a portable, self-contained device. The microfluidic chip houses microchannels that are 20-40 micron deep, 100 micron wide and a few centimeters long (similar in scale to human hair). Numerous sample manipulation steps are required to complete the detailed analysis.

photo of two fingers hold a small microfluidic chip and graph showing rising fluorescence as MMP-8 concentration increases

Microfluidic chips allows rapid and sensitive detection of salivary markers of periodontal disease

Seamless integration of these steps – sample preparation, mixing, and analytical assays – is made possible through a photopolymerization fabrication process, adapted from the semiconductor industry, which allows us to engineer cast-in-place nanoporous polymeric gels within the tiny channels. Gel polymerization is rapid and confined to microchannels exposed to light through the use of photoinitiators – an approach that yields customizable gel properties. Our photolithographic approach allows us to create a thin nanoporous membrane, having specific size-exclusion properties, localized in a “sample loading region” of the device. A multi-step process is easily performed at the membrane – saliva clean-up, concentration, and mixing with reagents. During the rapid mixing step, disease biomarkers present in the sample bind to fluorescently-labeled antibody probes. Polymeric elements with larger pores (separation gel) are located adjacent to the size exclusion membrane, thus allowing us to perform electrophoretically-driven molecular sieving of proteins that were retained at the size-exclusion membrane. As the now highly concentrated, purified mixture is transported through the nanoporous separation gel, molecules are separated based on their size and electrical charge. If biomarkers for the disease are present in the saliva, the lab-on-a-chip analysis will detect, via a photomultiplier tube, the presence of biomarkers, now bound to the fluorescent antibodies. IMPOD then rapidly determines the amount of biomarker present in the saliva.

The beauty and power of our approach is the seamless integration of all necessary steps — sample loading, mixing of sample with fluorescent detection antibody, concentration (to improve detection limit) and electrophoretic separation of bound and unbound antibody – into one instrument making operation “hands-free”. The entire device, including the microfluidic core, photomultiplier, and electronics, is a hand-held package that weighs less than five pounds, thus providing the features required for point-of-care applications: sensitivity, portability, and the ability to run tests quickly.

For more information

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Proteomics

Proteins are where the action is in a cell – they are the active end products of the genome. As such, direct, large-scale characterization of cellular proteins is key to understanding many biological processes where important information about protein expression, activity, or modification cannot be obtained at the DNA/RNA level. We have a number of efforts ongoing as listed below:

photo depicting two dimensional gel electrophoresis of soluble proteins in sulfate-reducing microbe Desulfovibrio vulgaris

Two-dimensional gel electrophoresis of soluble proteins in sulfate-reducing microbe Desulfovibrio vulgaris (photo courtesy Rajat Sapra)

  1. Proteomic survey using 2-dimensional gel electrophoresis/mass spectrometry—we have used shot-gun proteomics to identify proteins differentially expressed in cells upon perturbation such as heat-shock or challenge by a toxin.
  2. Identifying protein-protein interactions- Using affinity pull-downs and LC-MS-MS, we are identifying protein complexes in bacterial and mammalian cells.
  3. Protein structure determination using mass spectrometry—Using intra-protein chemical crosslinking followed by mass spectrometric identification of crosslinked amino acids, we have developed a technique to determine 3-D structure of proteins that are hard to crystallize such as integral membrane proteins.

We maintain state of the art equipment for proteomics research including 3 mass spectrometers (Applied Biosystems Voyager DE PRO, Waters QTOF Ultima, Thermo LCQ), liquid chromatography capability (HP1100, Waters CapLC, Eksigent nano2DLC), and high throughput sample preparation robotics (Typhoon gel imager, Ettan spot picker, Genomic Solutions ProPrep).

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Microfluidics

SDS-PAGE in a chip

SDS-PAGE in a chip

High-throughput experimental biology increasingly demands large numbers of experiments performed in a shorter period of time. Microfluidics or “Lab-on-a-Chip” devices are an enabling technology that allows researchers to study complex biological problems that would be difficult, tedious, labor-intensive, or otherwise cost-prohibitive using conventional bench-scale techniques. Microfludics offers high sensitivity with inherently low reagent consumption, and can be automated to perform a variety of complex, multi-step analyses with only minimal user intervention.

The Biosystems Research department has developed expertise in several areas of microfluidics, with an emphasis on applying proven lab-on-a-chip technology to solving relevant biological problems. Our strength in fundamental biology provides important insight and direction in choosing problems and developing techniques that will make an impact in the way researchers study biology. Our collaborations with the Microfluidics and Microsystems Engineering departments (8324 and 8125) provide additional strength in developing robust, miniaturized, user-friendly, automated systems for studying complex biological problems.

Areas of expertise in microfluidics within the Biosystems Research department include:

  • Photopatterning of functional elements, including sieving gels for electrophoresis, size-exclusion and dialysis membranes, and porous polymer monoliths for chromatography.
  • Microchip electrophoretic immunoassays for detection of biotoxins, biomarkers, cytokines, and other proteins in complex samples, with on-chip preconcentration and mixing of reagents.
  • Concentration, staining, and analysis of live cells on chip (part of MISL Grand Challenge).
  • Micropreparative extraction of proteins from complex biological samples.
  • Interfacing microfluidic separations with advanced detection technologies, including confocal microscopy, hyperspectral imaging (8332) and time-resolved multispectral single-molecule detection (8353).