Publications by authors named "Christopher Lausted"

24 Publications

  • Page 1 of 1

Single-cell analysis of erythropoiesis in Rpl11 haploinsufficient mice reveals insight into the pathogenesis of Diamond-Blackfan anemia.

Exp Hematol 2021 Feb 22. Epub 2021 Feb 22.

Department of Medicine, Division of Hematology, University of Washington, Seattle, WA.

Rpl11 haploinsufficient mice develop a macrocytic anemia similar to patients with DBA. Here, we fully characterize this model from clinical and pathophysiological perspectives. Early erythroid precursors have increased heme content and high cytoplasmic reactive oxygen species, impairing erythroid differentiation at the colony-forming unit-erythroid (CFU-E)/proerythroblast stage and subsequently. Using single-cell analyses that link a cell's surface protein expression to its total transcriptome and unbiased analyses, we found GATA1, GATA1 target gene, and mitotic spindle pathway gene transcription were the pathways that decreased the most. Expression of ribosome protein and globin genes was amplified. These changes, as well as the other transcriptional changes that were identified, closely resemble findings in mice that lack the heme export protein FLVCR and, thus, suggest that heme excess and toxicity are the primary drivers of the macrocytic anemia. Consistent with this, treating Rpl11 haploinsufficient mice with corticosteroids increased the numbers of earliest erythroblasts but failed to overcome heme toxicity and improve the anemia. Rpl11 haploinsufficient mice uniquely upregulated mitochondrial genes, p53 and CDKN1A pathway genes, and DNA damage checkpoint genes, which should contribute further to erythroid marrow failure. Together our data establish Rpl11 haploinsufficient mice as an excellent model of DBA that can be used to study DBA pathogenesis and test novel therapies.
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http://dx.doi.org/10.1016/j.exphem.2021.02.010DOI Listing
February 2021

Modulation of Immune Checkpoints by Chemotherapy in Human Colorectal Liver Metastases.

Cell Rep Med 2020 Dec 22;1(9):100160. Epub 2020 Dec 22.

Institute for Systems Biology, Seattle, WA, USA.

Metastatic colorectal cancer (CRC) is a major cause of cancer-related death, and incidence is rising in younger populations (younger than 50 years). Current chemotherapies can achieve response rates above 50%, but immunotherapies have limited value for patients with microsatellite-stable (MSS) cancers. The present study investigates the impact of chemotherapy on the tumor immune microenvironment. We treat human liver metastases slices with 5-fluorouracil (5-FU) plus either irinotecan or oxaliplatin, then perform single-cell transcriptome analyses. Results from eight cases reveal two cellular subtypes with divergent responses to chemotherapy. Susceptible tumors are characterized by a stemness signature, an activated interferon pathway, and suppression of PD-1 ligands in response to 5-FU+irinotecan. Conversely, immune checkpoint TIM-3 ligands are maintained or upregulated by chemotherapy in CRC with an enterocyte-like signature, and combining chemotherapy with TIM-3 blockade leads to synergistic tumor killing. Our analyses highlight chemomodulation of the immune microenvironment and provide a framework for combined chemo-immunotherapies.
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http://dx.doi.org/10.1016/j.xcrm.2020.100160DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7762777PMC
December 2020

Multi-Omics Resolves a Sharp Disease-State Shift between Mild and Moderate COVID-19.

Cell 2020 12 28;183(6):1479-1495.e20. Epub 2020 Oct 28.

Institute for Systems Biology, Seattle, WA 98109, USA; Department of Bioengineering, University of Washington, Seattle, WA 98105, USA. Electronic address:

We present an integrated analysis of the clinical measurements, immune cells, and plasma multi-omics of 139 COVID-19 patients representing all levels of disease severity, from serial blood draws collected during the first week of infection following diagnosis. We identify a major shift between mild and moderate disease, at which point elevated inflammatory signaling is accompanied by the loss of specific classes of metabolites and metabolic processes. Within this stressed plasma environment at moderate disease, multiple unusual immune cell phenotypes emerge and amplify with increasing disease severity. We condensed over 120,000 immune features into a single axis to capture how different immune cell classes coordinate in response to SARS-CoV-2. This immune-response axis independently aligns with the major plasma composition changes, with clinical metrics of blood clotting, and with the sharp transition between mild and moderate disease. This study suggests that moderate disease may provide the most effective setting for therapeutic intervention.
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http://dx.doi.org/10.1016/j.cell.2020.10.037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7598382PMC
December 2020

Multiomic Immunophenotyping of COVID-19 Patients Reveals Early Infection Trajectories.

bioRxiv 2020 Jul 31. Epub 2020 Jul 31.

Host immune responses play central roles in controlling SARS-CoV2 infection, yet remain incompletely characterized and understood. Here, we present a comprehensive immune response map spanning 454 proteins and 847 metabolites in plasma integrated with single-cell multi-omic assays of PBMCs in which whole transcriptome, 192 surface proteins, and T and B cell receptor sequence were co-analyzed within the context of clinical measures from 50 COVID19 patient samples. Our study reveals novel cellular subpopulations, such as proliferative exhausted CD8 and CD4 T cells, and cytotoxic CD4 T cells, that may be features of severe COVID-19 infection. We condensed over 1 million immune features into a single immune response axis that independently aligns with many clinical features and is also strongly associated with disease severity. Our study represents an important resource towards understanding the heterogeneous immune responses of COVID-19 patients and may provide key information for informing therapeutic development.
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http://dx.doi.org/10.1101/2020.07.27.224063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7402042PMC
July 2020

Single-cell analyses demonstrate that a heme-GATA1 feedback loop regulates red cell differentiation.

Blood 2019 01 10;133(5):457-469. Epub 2018 Dec 10.

Department of Medicine, Division of Hematology, University of Washington, Seattle, WA; and.

Erythropoiesis is the complex, dynamic, and tightly regulated process that generates all mature red blood cells. To understand this process, we mapped the developmental trajectories of progenitors from wild-type, erythropoietin-treated, and -deleted mice at single-cell resolution. Importantly, we linked the quantity of each cell's surface proteins to its total transcriptome, which is a novel method. Deletion of results in high levels of intracellular heme, allowing us to identify heme-regulated circuitry. Our studies demonstrate that in early erythroid cells (CD71Ter119), heme increases ribosomal protein transcripts, suggesting that heme, in addition to upregulating globin transcription and translation, guarantees ample ribosomes for globin synthesis. In later erythroid cells (CD71Ter119), heme decreases GATA1, GATA1-target gene, and mitotic spindle gene expression. These changes occur quickly. For example, in confirmatory studies using human marrow erythroid cells, ribosomal protein transcripts and proteins increase, and GATA1 transcript and protein decrease, within 15 to 30 minutes of amplifying endogenous heme synthesis with aminolevulinic acid. Because GATA1 initiates heme synthesis, GATA1 and heme together direct red cell maturation, and heme stops GATA1 synthesis, our observations reveal a GATA1-heme autoregulatory loop and implicate GATA1 and heme as the comaster regulators of the normal erythroid differentiation program. In addition, as excessive heme could amplify ribosomal protein imbalance, prematurely lower GATA1, and impede mitosis, these data may help explain the ineffective (early termination of) erythropoiesis in Diamond Blackfan anemia and del(5q) myelodysplasia, disorders with excessive heme in colony-forming unit-erythroid/proerythroblasts, explain why these anemias are macrocytic, and show why children with GATA1 mutations have DBA-like clinical phenotypes.
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http://dx.doi.org/10.1182/blood-2018-05-850412DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6356983PMC
January 2019

Novel metrics for quantifying bacterial genome composition skews.

BMC Genomics 2018 Jul 11;19(1):528. Epub 2018 Jul 11.

Institute for Systems Biology, 401 Terry Ave N, Seattle, WA, 98109, USA.

Background: Bacterial genomes have characteristic compositional skews, which are differences in nucleotide frequency between the leading and lagging DNA strands across a segment of a genome. It is thought that these strand asymmetries arise as a result of mutational biases and selective constraints, particularly for energy efficiency. Analysis of compositional skews in a diverse set of bacteria provides a comparative context in which mutational and selective environmental constraints can be studied. These analyses typically require finished and well-annotated genomic sequences.

Results: We present three novel metrics for examining genome composition skews; all three metrics can be computed for unfinished or partially-annotated genomes. The first two metrics, (dot-skew and cross-skew) depend on sequence and gene annotation of a single genome, while the third metric (residual skew) highlights unusual genomes by subtracting a GC content-based model of a library of genome sequences. We applied these metrics to 7738 available bacterial genomes, including partial drafts, and identified outlier species. A phylogenetically diverse set of these outliers (i.e., Borrelia, Ehrlichia, Kinetoplastibacterium, and Phytoplasma) display similar skew patterns but share lifestyle characteristics, such as intracellularity and biosynthetic dependence on their hosts.

Conclusions: Our novel metrics appear to reflect the effects of biosynthetic constraints and adaptations to life within one or more hosts on genome composition. We provide results for each analyzed genome, software and interactive visualizations at http://db.systemsbiology.net/gestalt/ skew_metrics .
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http://dx.doi.org/10.1186/s12864-018-4913-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6042203PMC
July 2018

Whole genome sequence and comparative analysis of Borrelia burgdorferi MM1.

PLoS One 2018 11;13(6):e0198135. Epub 2018 Jun 11.

Institute for Systems Biology, Seattle, Washington, United States of America.

Lyme disease is caused by spirochaetes of the Borrelia burgdorferi sensu lato genospecies. Complete genome assemblies are available for fewer than ten strains of Borrelia burgdorferi sensu stricto, the primary cause of Lyme disease in North America. MM1 is a sensu stricto strain originally isolated in the midwestern United States. Aside from a small number of genes, the complete genome sequence of this strain has not been reported. Here we present the complete genome sequence of MM1 in relation to other sensu stricto strains and in terms of its Multi Locus Sequence Typing. Our results indicate that MM1 is a new sequence type which contains a conserved main chromosome and 15 plasmids. Our results include the first contiguous 28.5 kb assembly of lp28-8, a linear plasmid carrying the vls antigenic variation system, from a Borrelia burgdorferi sensu stricto strain.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0198135PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5995427PMC
January 2019

A wellness study of 108 individuals using personal, dense, dynamic data clouds.

Nat Biotechnol 2017 Aug 17;35(8):747-756. Epub 2017 Jul 17.

Institute for Systems Biology, Seattle, Washington, USA.

Personal data for 108 individuals were collected during a 9-month period, including whole genome sequences; clinical tests, metabolomes, proteomes, and microbiomes at three time points; and daily activity tracking. Using all of these data, we generated a correlation network that revealed communities of related analytes associated with physiology and disease. Connectivity within analyte communities enabled the identification of known and candidate biomarkers (e.g., gamma-glutamyltyrosine was densely interconnected with clinical analytes for cardiometabolic disease). We calculated polygenic scores from genome-wide association studies (GWAS) for 127 traits and diseases, and used these to discover molecular correlates of polygenic risk (e.g., genetic risk for inflammatory bowel disease was negatively correlated with plasma cystine). Finally, behavioral coaching informed by personal data helped participants to improve clinical biomarkers. Our results show that measurement of personal data clouds over time can improve our understanding of health and disease, including early transitions to disease states.
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http://dx.doi.org/10.1038/nbt.3870DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5568837PMC
August 2017

Microarray based screening of peptide nano probes for HER2 positive tumor.

Anal Chem 2015 Aug 6;87(16):8367-72. Epub 2015 Aug 6.

§Institute for Systems Biology, 401 Terry Avenue North, Seattle, Washington 98109, United States.

Peptides are excellent biointerface molecules and diagnostic probes with many advantages such as good penetration, short turnover time, and low cost. We report here an efficient peptide screening strategy based on in situ single bead sequencing on a microarray. Two novel peptides YLFFVFER (H6) and KLRLEWNR (H10) specifically binding to the tumor biomarker human epidermal growth factor receptor 2 (HER2) with aKD of 10(-8) M were obtained from a 10(5) library. Conjugated to nanoparticles, both the H6 and H10 probes showed specific accumulation in HER2-positive tumor tissues in xenografted mice by in vivo imaging.
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http://dx.doi.org/10.1021/acs.analchem.5b01588DOI Listing
August 2015

Plain silver surface plasmon resonance for microarray application.

Anal Chem 2015 Feb 14;87(3):1466-9. Epub 2015 Jan 14.

National Center for NanoScience and Technology , Beijing 100190, People's Republic of China.

The application scope of surface plasmon resonance (SPR) and SPR imaging (SPRi) is rapidly growing, and tools such as high-performance and low-cost slides could enable more rapid growth of the field. We describe herein a novel silver slide, addressing the inherent instability of plain silver structure by improving adhesion between the glass substrate and the silver layer with a thin buffer layer of gold. Covered by a self-assembled monolayer (SAM) only, SPR characteristics of the slide remain steady for more than 3 months under regular storage. In a bioassay, the slide substantiates the predicted nearly 100% sensitivity improvement over gold slides and exhibits exceptional performance stability as determined by sensitivity and resolution measurements during the extended 40,000 s multicycle experiment. We demonstrate the suitability of this new slide for large-area SPRi, describing analysis results from a 1 296-ligand protein microarray. We predict this slide structure will provide a stable, high-sensitivity solution for high-throughput SPRi applications and other surface analysis platforms.
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http://dx.doi.org/10.1021/ac504110tDOI Listing
February 2015

Rapid screening of peptide probes through in situ single-bead sequencing microarray.

Anal Chem 2014 Dec 14;86(23):11854-9. Epub 2014 Nov 14.

CAS Key Laboratory for Biomedical Effects of Nanomaterials & Nanosafety, National Center for Nanoscience and Technology of China , Beijing 100190, China.

Peptide ligands as targeting probes for in vivo imaging and drug delivery have attracted great interest in the biomedical community. However, high affinity and specificity screening of large peptide libraries remains a tedious process. Here, we report a continuous-flow microfluidic method for one-bead-one-compound (OBOC) combinatorial peptide library screening. We screened a library with 2 × 10(5) peptide beads within 4 h and discovered 140 noncanonical peptide hits targeting the tumor marker, aminopeptidase N (APN). Using the Clustal algorithm, we identified the conserved sequence Tyr-XX-Tyr in the N terminal. We demonstrated that the novel sequence YVEYHLC peptides have both nanomolar affinity and high specificity for APN in ex vivo and in vivo models. We envision that the successful demonstration of this integrated novel nanotechnology for peptide screening and identification open a new avenue for rapid discovery of new peptide-based reagents for disease diagnostics and therapeutics.
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http://dx.doi.org/10.1021/ac503454zDOI Listing
December 2014

Betaine homocysteine methyltransferase (BHMT) as a specific and sensitive blood marker for acute liver injury.

Biomarkers 2014 Nov 21;19(7):578-84. Epub 2014 Aug 21.

National Center for Nanoscience and Technology , Beijing , P.R. China .

We developed a high-performance ELISA assay and measured serum BHMT levels in healthy individuals and patients with acute liver injury (ALI). The detection range of this ELISA assay was from 1.56 to 100 ng/ml. BHMT levels are significantly higher in ALI groups. In the healthy group (n = 244), the median value (interquartile range, IQR 0-56.40) was 1.83 ng/ml. In the ALI group (n = 42), the median value of BHMT was 748.48 ng/ml (IQR, 0-51095.92). ROC curve analysis demonstrated good sensitivity (0.86) and specificity (0.98). In addition, in five ALI cases with time course samples available, BHMT and ALT both followed the "rise and fall" temporal pattern with the disease progression. However, the slopes of BHMT curves were steeper than ALT curves. And in three out of the five cases, BHMT levels peaked 1 day earlier than ALT levels be a sensitive marker with good prognostic value.
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http://dx.doi.org/10.3109/1354750X.2014.951880DOI Listing
November 2014

Label-free quantitative detection of tumor-derived exosomes through surface plasmon resonance imaging.

Anal Chem 2014 Sep 11;86(17):8857-64. Epub 2014 Aug 11.

National Center for Nanoscience and Technology , No. 11, Beiyitiao Zhongguancun, Beijing 100190, P. R. China.

Exosomes are endosome-derived membrane vesicles carrying proteins and nucleic acids that are involved in cellular functions such as intercellular communication, protein and RNA secretion, and antigen presentation. Therefore, exosomes serve as potential biomarkers for many diseases including cancer. Because exosomes are difficult to enrich or purify from biofluids, quantification of exosomes is tedious and inaccurate. Here, we present a real-time, label-free, and quantitative method to detect and characterize tumor-derived exosomes without enrichment or purification. Utilizing surface plasmon resonance imaging (SPRi) in combination with antibody microarrays specific to the extracellular domains of exosome membrane proteins, exosomes in tumor cell culture medium can be quantitatively detected. We found a positive correlation between the metastatic potential of tumor cell lines and exosome secretion. This method provides an easy, efficient, and novel way to detect exosome secretion and thus an avenue toward the diagnosis and prognosis prediction of cancer.
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http://dx.doi.org/10.1021/ac5023056DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4151789PMC
September 2014

Quantitative liver-specific protein fingerprint in blood: a signature for hepatotoxicity.

Theranostics 2014 14;4(2):215-28. Epub 2014 Jan 14.

2. Institute for Systems Biology, 401 Terry Avenue N, Seattle, Washington 98109, USA.

We discuss here a new approach to detecting hepatotoxicity by employing concentration changes of liver-specific blood proteins during disease progression. These proteins are capable of assessing the behaviors of their cognate liver biological networks for toxicity or disease perturbations. Blood biomarkers are highly desirable diagnostics as blood is easily accessible and baths virtually all organs. Fifteen liver-specific blood proteins were identified as markers of acetaminophen (APAP)-induced hepatotoxicity using three proteomic technologies: label-free antibody microarrays, quantitative immunoblotting, and targeted iTRAQ mass spectrometry. Liver-specific blood proteins produced a toxicity signature of eleven elevated and four attenuated blood protein levels. These blood protein perturbations begin to provide a systems view of key mechanistic features of APAP-induced liver injury relating to glutathione and S-adenosyl-L-methionine (SAMe) depletion, mitochondrial dysfunction, and liver responses to the stress. Two markers, elevated membrane-bound catechol-O-methyltransferase (MB-COMT) and attenuated retinol binding protein 4 (RBP4), report hepatic injury significantly earlier than the current gold standard liver biomarker, alanine transaminase (ALT). These biomarkers were perturbed prior to onset of irreversible liver injury. Ideal markers should be applicable for both rodent model studies and human clinical trials. Five of these mouse liver-specific blood markers had human orthologs that were also found to be responsive to human hepatotoxicity. This panel of liver-specific proteins has the potential to effectively identify the early toxicity onset, the nature and extent of liver injury and report on some of the APAP-perturbed liver networks.
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http://dx.doi.org/10.7150/thno.7868DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3900804PMC
September 2014

Systems approach to neurodegenerative disease biomarker discovery.

Annu Rev Pharmacol Toxicol 2014 23;54:457-81. Epub 2013 Oct 23.

Institute for Systems Biology, Seattle, Washington 98109; email: , , , , , ,

Biomarkers are essential for performing early diagnosis, monitoring neurodegenerative disease progression, gauging responses to therapies, and stratifying neurodegenerative diseases into their different subtypes. A wide range of molecular markers are under investigation in tissues and biofluids as well as through imaging; moreover, many are prominent proteins present in cerebrospinal fluid. However, in more frequently and easily collected fluids such as plasma, these proteins show only a modest correlation with disease and thus lack the necessary sensitivity or specificity for clinical use. High-throughput and quantitative proteomic technologies and systems-driven approaches to biofluid analysis are now being utilized in the search for better biomarkers. Biomarker discovery involves many critical steps including study design, sample preparation, protein and peptide separation and identification, and bioinformatics and data integration issues that must be carefully controlled before independent confirmation and validation. In this review, we summarize current proteomic and nucleic acid technologies involved in the discovery of biomarkers of neurodegenerative diseases, particularly Alzheimer's, Parkinson's, Huntington's, and prion diseases.
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http://dx.doi.org/10.1146/annurev-pharmtox-011613-135928DOI Listing
September 2014

An automated Teflon microfluidic peptide synthesizer.

Lab Chip 2013 Sep 8;13(17):3347-50. Epub 2013 Jul 8.

National Center for Nanoscience and Technology, No.11 ZhongGuanCun BeiYiTiao, 100190 Beijing, PR China.

We present a microfluidic synthesizer made entirely of Teflon material for solid phase peptide synthesis (SPPS). Solvent-resistant perfluoroalkoxy (PFA) was used to construct chip-sized devices featuring multiple tri-layer pneumatic microvalves. Using these devices, model peptides were automatically synthesized and cleaved in situ in a continuous-flow manner. The total coupling and cleavage time was significantly reduced compared to conventional bulk reactors. The synthesis of a decapeptide, for instance, took less than 6 h using our device while it usually takes more than three days using conventional reactors.
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http://dx.doi.org/10.1039/c3lc50632kDOI Listing
September 2013

Label-free detection with surface plasmon resonance imaging.

Methods Mol Biol 2011 ;723:321-33

Institute for Systems Biology, Seattle, WA, USA.

Sensors based on surface plasmon resonance have demonstrated, over the last 2 decades, to be an effective method of studying biomolecular interactions without the need for labeling. Recently, it has been adapted to high-throughput use for imaging microarray binding in real time. This provides a promising platform - a label-free protein microarray system - for the study of disease. In this example, antibody microarrays are used to efficiently profile the secretion of proteins from a cell line exposed to varying concentrations of a toxic compound.
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http://dx.doi.org/10.1007/978-1-61779-043-0_20DOI Listing
June 2011

Parallel microfluidic surface plasmon resonance imaging arrays.

Lab Chip 2010 Mar 6;10(5):581-8. Epub 2010 Jan 6.

Michael Smith Laboratories, University of British Columbia, Vancouver, BC, Canada V6T 1Z4.

Surface plasmon resonance imaging (SPRi) is a label-free technique used for the quantitation of binding affinities and concentrations for a wide variety of target molecules. Although SPRi is capable of determining binding constants for multiple ligands in parallel, current commercial instruments are limited to a single analyte stream on multiple ligand spots. Measurement of binding kinetics requires the serial introduction of different analyte concentrations; such repeated experiments are conducted manually and are therefore time-intensive. To address these challenges, we have developed an integrated microfluidic array using soft lithography techniques for high-throughput SPRi-based detection and determination of binding affinities of antibodies against protein targets. The device consists of 264 element-addressable chambers isolated by microvalves. The resulting 700 pL chamber volumes, combined with a serial dilution network for simultaneous interrogation of up to six different analyte concentrations, allow for further speeding detection times. To test for device performance, human alpha-thrombin was immobilized on the sensor surface and anti-human alpha-thrombin IgG was injected across the surface at different concentrations. The equilibrium dissociation constant was determined to be 5.0 +/- 1.9 nM, which agrees well with values reported in the literature. The interrogation of multiple ligands to multiple analytes in a single device was also investigated and samples were recovered with no cross-contamination. Since each chamber can be addressed independently, this array is capable of interrogating binding events from up to 264 different immobilized ligands against multiple analytes in a single experiment. The development of high-throughput protein analytic measurements is a critical technology for systems approaches to biology and medicine.
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http://dx.doi.org/10.1039/b920589fDOI Listing
March 2010

SPR imaging for high throughput, label-free interaction analysis.

Comb Chem High Throughput Screen 2009 Sep 1;12(8):741-51. Epub 2009 Sep 1.

Institute for Systems Biology, Seattle, Washington 98103, USA.

Surface plasmon resonance (SPR) sensors have proven themselves over the last 20 years to be an effective method to study biomolecular binding and kinetics without the use of labeling. More recently, the approach has been adapted to high-throughput use with the imaging of SPR-active microarrays. This is an excellent tool for monitoring microarray binding in real-time where the microarray probes and targets can include a wide range of molecules. DNA, RNA, antibodies, enzymes, and a range of other proteins have been arrayed and quantitatively analyzed.
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http://dx.doi.org/10.2174/138620709789104933DOI Listing
September 2009

Quantitative serum proteomics from surface plasmon resonance imaging.

Mol Cell Proteomics 2008 Dec 3;7(12):2464-74. Epub 2008 Aug 3.

Institute for Systems Biology, Seattle, Washington 98103, USA.

The detection and quantification of specific proteins in complex mixtures is a major challenge for proteomics. For example, the development of disease-related biomarker panels will require fast and efficient methods for obtaining multiparameter protein profiles. We established a high throughput, label-free method for analyzing serum using surface plasmon resonance imaging of antibody microarrays. Microarrays were fabricated using standard pin spotting on bare gold substrates, and samples were applied for binding analysis using a camera-based surface plasmon resonance system. We validated the system by measuring the concentrations of four serum proteins using part of a 792-feature microarray. Transferrin concentrations were measured to be 2.1 mg/ml in human serum and 1.2 mg/ml in murine serum, which closely matched ELISA determinations of 2.6 and 1.2 mg/ml, respectively. In agreement with expected values, human and mouse albumin levels were measured to be 24.3 and 23.6 mg/ml, respectively. The lower limits of detection for the four measurements ranged from 14 to 58 ng/ml or 175 to 755 pm. Where purified target proteins are not available for calibration, the microarrays can be used for relative protein quantification. We used the antibody microarray to compare the serum protein profiles from three liver cancer patients and three non-liver cancer patients. Hierarchical clustering of the serum protein levels clearly distinguished two distinct profiles. Thirty-nine significant protein changes were detected (p < 0.05), 10 of which have been observed previously in serum. alpha-Fetoprotein, a known liver cancer marker, was observed to increase. These results demonstrate the feasibility of this high throughput approach for both absolute and relative protein expression profiling.
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http://dx.doi.org/10.1074/mcp.M800121-MCP200DOI Listing
December 2008

Printing your own inkjet microarrays.

Methods Enzymol 2006 ;410:168-89

Institute for Systems Biology, Seattle, Washington, USA.

DNA arrays are now the tools of choice for high-throughput DNA/RNA analysis. While many technologies exist for mass-producing arrays, there are just a few ways to economically produce small batches of custom oligonucleotide arrays for prototyping experiments and specialized applications. Inkjet printing, adapted from the world of office electronics to the world of molecular biology, is one such method. With programmable oligonucleotide synthesizers, scientists can prototype DNA array assays quickly and inexpensively. A benchtop inkjet arrayer-nicknamed POSAM-can be built by most skilled molecular biology laboratories. Inkjet arrays can fulfill the changing needs of those studying the complex network of relationships in systems biology.
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http://dx.doi.org/10.1016/S0076-6879(06)10008-7DOI Listing
December 2006

Maximum static inspiratory and expiratory pressures with different lung volumes.

Biomed Eng Online 2006 May 5;5:29. Epub 2006 May 5.

The Institute for Systems Biology, 1441 North 34th Street, Seattle, WA 98103, USA.

Background: Maximum pressures developed by the respiratory muscles can indicate the health of the respiratory system, help to determine maximum respiratory flow rates, and contribute to respiratory power development. Past measurements of maximum pressures have been found to be inadequate for inclusion in some exercise models involving respiration.

Methods: Maximum inspiratory and expiratory airway pressures were measured over a range of lung volumes in 29 female and 19 male adults. A commercial bell spirometry system was programmed to occlude airflow at nine target lung volumes ranging from 10% to 90% of vital capacity.

Results: In women, maximum expiratory pressure increased with volume from 39 to 61 cmH2O and maximum inspiratory pressure decreased with volume from 66 to 28 cmH2O. In men, maximum expiratory pressure increased with volume from 63 to 97 cmH2O and maximum inspiratory pressure decreased with volume from 97 to 39 cmH2O. Equations describing pressures for both sexes are: Pe/Pmax = 0.1426 Ln( %VC) + 0.3402 R2 = 0.95 Pi/Pmax = 0.234 Ln(100 - %VC) - 0.0828 R2 = 0.96

Conclusion: These results were found to be consistent with values and trends obtained by other authors. Regression equations may be suitable for respiratory mechanics models.
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http://dx.doi.org/10.1186/1475-925X-5-29DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1501025PMC
May 2006

POSaM: a fast, flexible, open-source, inkjet oligonucleotide synthesizer and microarrayer.

Genome Biol 2004 27;5(8):R58. Epub 2004 Jul 27.

The Institute for Systems Biology, 1441 North 34th Street, Seattle, WA 98103, USA.

DNA arrays are valuable tools in molecular biology laboratories. Their rapid acceptance was aided by the release of plans for a pin-spotting microarrayer by researchers at Stanford. Inkjet microarraying is a flexible, complementary technique that allows the synthesis of arrays of any oligonucleotide sequences de novo. We describe here an open-source inkjet arrayer capable of rapidly producing sets of unique 9,800-feature arrays.
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http://dx.doi.org/10.1186/gb-2004-5-8-r58DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC507883PMC
February 2005

Effects of caffeine on performance of low intensity tasks.

Percept Mot Skills 2002 Apr;94(2):521-32

Department of Biological Resources Engineering, University of Maryland, College Park 20742-2315, USA.

31 college age men and women who consume less than three caffeinated beverages per week agreed to participate as subjects in research on the effects of acute caffeine intake on low intensity task performance. All subjects performed two randomly administered test conditions: (1) caffeine (5 mg/kg) and (2) placebo on separate visits following an initial 1-hr. orientation visit. Subjects were administered the beverage 30 min. prior to performing 12 separate tests assessing basic mathematics, simple response, logical reasoning, hand-eye coordination, and spatial and assembly skills. The Spielberger State Anxiety test was administered immediately after consuming the test beverage and once again at posttest. Analysis showed that caffeine did not significantly affect performance on all tests with the exception of the peripheral awareness (hand-eye coordination) test on which performance was higher after ingesting caffeine. The placebo treatment produced no effect on state anxiety, which contrasted with a significant rise in anxiety after caffeine consumption. State anxiety values were significantly greater after caffeine treatment relative to the placebo at pretest, and this difference persisted at posttest. These results demonstrated that the dose of caffeine increased scores on state anxiety for individuals who consumed less than three caffeinated beverages weekly but had very little effect on performance of low intensity tasks, except for a hand-eye coordination test involving peripheral awareness. Perhaps longer continuous performance of more demanding tasks would be more sensitive.
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http://dx.doi.org/10.2466/pms.2002.94.2.521DOI Listing
April 2002