Journal Articles

  • CytoSPADE: high-performance analysis and visualization of high-dimensional cytometry data BIOINFORMATICS Linderman, M. D., Bjornson, Z., Simonds, E. F., Qiu, P., Bruggner, R. V., Sheode, K., Meng, T. H., Plevritis, S. K., Nolan, G. P. 2012; 28 (18): 2400-2401


    MOTIVATION: Recent advances in flow cytometry enable simultaneous single-cell measurement of 30+ surface and intracellular proteins. CytoSPADE is a high-performance implementation of an interface for the Spanning-tree Progression Analysis of Density-normalized Events algorithm for tree-based analysis and visualization of this high-dimensional cytometry data. AVAILABILITY: Source code and binaries are freely available at and via Bioconductor version 2.10 onwards for Linux, OSX and Windows. CytoSPADE is implemented in R, C++ and Java. CONTACT: SUPPLEMENTARY INFORMATION: Additional documentation available at

    View details for DOI 10.1093/bioinformatics/bts425

    View details for Web of Science ID 000308532300067

    View details for PubMedID 22782546

  • Multiplexed mass cytometry profiling of cellular states perturbed by small-molecule regulators NATURE BIOTECHNOLOGY Bodenmiller, B., Zunder, E. R., Finck, R., Chen, T. J., Savig, E. S., Bruggner, R. V., Simonds, E. F., Bendall, S. C., Sachs, K., Krutzik, P. O., Nolan, G. P. 2012; 30 (9): 858-U89


    Mass cytometry facilitates high-dimensional, quantitative analysis of the effects of bioactive molecules on human samples at single-cell resolution, but instruments process only one sample at a time. Here we describe mass-tag cellular barcoding (MCB), which increases mass cytometry throughput by using n metal ion tags to multiplex up to 2n samples. We used seven tags to multiplex an entire 96-well plate, and applied MCB to characterize human peripheral blood mononuclear cell (PBMC) signaling dynamics and cell-to-cell communication, signaling variability between PBMCs from eight human donors, and the effects of 27 inhibitors on this system. For each inhibitor, we measured 14 phosphorylation sites in 14 PBMC types at 96 conditions, resulting in 18,816 quantified phosphorylation levels from each multiplexed sample. This high-dimensional, systems-level inquiry allowed analysis across cell-type and signaling space, reclassified inhibitors and revealed off-target effects. High-content, high-throughput screening with MCB should be useful for drug discovery, preclinical testing and mechanistic investigation of human disease.

    View details for DOI 10.1038/nbt.2317

    View details for Web of Science ID 000308705700020

    View details for PubMedID 22902532

  • Extracting a cellular hierarchy from high-dimensional cytometry data with SPADE NATURE BIOTECHNOLOGY Qiu, P., Simonds, E. F., Bendall, S. C., Gibbs, K. D., Bruggner, R. V., Linderman, M. D., Sachs, K., Nolan, G. P., Plevritis, S. K. 2011; 29 (10): 886-U181


    The ability to analyze multiple single-cell parameters is critical for understanding cellular heterogeneity. Despite recent advances in measurement technology, methods for analyzing high-dimensional single-cell data are often subjective, labor intensive and require prior knowledge of the biological system. To objectively uncover cellular heterogeneity from single-cell measurements, we present a versatile computational approach, spanning-tree progression analysis of density-normalized events (SPADE). We applied SPADE to flow cytometry data of mouse bone marrow and to mass cytometry data of human bone marrow. In both cases, SPADE organized cells in a hierarchy of related phenotypes that partially recapitulated well-described patterns of hematopoiesis. We demonstrate that SPADE is robust to measurement noise and to the choice of cellular markers. SPADE facilitates the analysis of cellular heterogeneity, the identification of cell types and comparison of functional markers in response to perturbations.

    View details for DOI 10.1038/nbt.1991

    View details for Web of Science ID 000296273000015

    View details for PubMedID 21964415

  • Single-Cell Mass Cytometry of Differential Immune and Drug Responses Across a Human Hematopoietic Continuum SCIENCE Bendall, S. C., Simonds, E. F., Qiu, P., Amir, E. D., Krutzik, P. O., Finck, R., Bruggner, R. V., Melamed, R., Trejo, A., Ornatsky, O. I., Balderas, R. S., Plevritis, S. K., Sachs, K., Pe'er, D., Tanner, S. D., Nolan, G. P. 2011; 332 (6030): 687-696


    Flow cytometry is an essential tool for dissecting the functional complexity of hematopoiesis. We used single-cell "mass cytometry" to examine healthy human bone marrow, measuring 34 parameters simultaneously in single cells (binding of 31 antibodies, viability, DNA content, and relative cell size). The signaling behavior of cell subsets spanning a defined hematopoietic hierarchy was monitored with 18 simultaneous markers of functional signaling states perturbed by a set of ex vivo stimuli and inhibitors. The data set allowed for an algorithmically driven assembly of related cell types defined by surface antigen expression, providing a superimposable map of cell signaling responses in combination with drug inhibition. Visualized in this manner, the analysis revealed previously unappreciated instances of both precise signaling responses that were bounded within conventionally defined cell subsets and more continuous phosphorylation responses that crossed cell population boundaries in unexpected manners yet tracked closely with cellular phenotype. Collectively, such single-cell analyses provide system-wide views of immune signaling in healthy human hematopoiesis, against which drug action and disease can be compared for mechanistic studies and pharmacologic intervention.

    View details for DOI 10.1126/science.1198704

    View details for Web of Science ID 000290265800035

    View details for PubMedID 21551058

  • Update of the Anopheles gambiae PEST genome assembly GENOME BIOLOGY Sharakhova, M. V., Hammond, M. P., Lobo, N. F., Krzywinski, J., Unger, M. F., Hillenmeyer, M. E., Bruggner, R. V., Birney, E., Collins, F. H. 2007; 8 (1)


    The genome of Anopheles gambiae, the major vector of malaria, was sequenced and assembled in 2002. This initial genome assembly and analysis made available to the scientific community was complicated by the presence of assembly issues, such as scaffolds with no chromosomal location, no sequence data for the Y chromosome, haplotype polymorphisms resulting in two different genome assemblies in limited regions and contaminating bacterial DNA.Polytene chromosome in situ hybridization with cDNA clones was used to place 15 unmapped scaffolds (sizes totaling 5.34 Mbp) in the pericentromeric regions of the chromosomes and oriented a further 9 scaffolds. Additional analysis by in situ hybridization of bacterial artificial chromosome (BAC) clones placed 1.32 Mbp (5 scaffolds) in the physical gaps between scaffolds on euchromatic parts of the chromosomes. The Y chromosome sequence information (0.18 Mbp) remains highly incomplete and fragmented among 55 short scaffolds. Analysis of BAC end sequences showed that 22 inter-scaffold gaps were spanned by BAC clones. Unmapped scaffolds were also aligned to the chromosome assemblies in silico, identifying regions totaling 8.18 Mbp (144 scaffolds) that are probably represented in the genome project by two alternative assemblies. An additional 3.53 Mbp of alternative assembly was identified within mapped scaffolds. Scaffolds comprising 1.97 Mbp (679 small scaffolds) were identified as probably derived from contaminating bacterial DNA. In total, about 33% of previously unmapped sequences were placed on the chromosomes.This study has used new approaches to improve the physical map and assembly of the A. gambiae genome.

    View details for DOI 10.1186/gb-2007-8-1-r5

    View details for Web of Science ID 000244821100011

    View details for PubMedID 17210077

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