In the last year, 39 researchers used the resources of the Laboratory of Cellular and Molecular Biology Microscopy Core. While most of the researchers come from the Laboratory of Cellular and Molecular Biology, the Core has been used by scientists from the Laboratory of Metabolism, the Section of Cellular and Developmental Biology in the Program in Genomics of Differentiation,the Cell and Cancer Biology Branch and The University of Maryland Department of Physics. Almost all of the Principal Investigators in the Laboratory of Cellular and Molecular Biology have projects that involve the Core facility. Dr. Lawrence Samelson uses Core resources for the project "Biochemical Basis of T Cell Activation". Researchers working with Dr. Carol Clayberger have used the facility for the projects "Regulation of RANTES Expression in T Lymphocytes" and "Function of Granulysin". The Core has been involved with the project "Cbl Proteins as Regulators of Tyrosine Kinase Signaling"from Dr. Stanley Lipkowitz. Dr. Carole Parent's projects, "Signaling Events Regulating Chemotaxis" and"Chemotactic Signals Reulating Human Neutrophil and Breast Metastatic Migration" use Core instruments. Dr. Paul Randazzo has made extensive use of the Core for the projects "Regulation of focal adhesions" and "Turnover of invadopodia". The Core facility has assisted Dr.Jeffry Rubin with the project "Casein Kinase 1 Delta in Wnt Signaling and Beyond". Dr. Ying Zhang uses Core instruments for the project "Molecular Mechanisms of TGF-beta Signaling Pathway". In addition, the Core facility has been used by personnel working with Principal Investigators from other groups including work with Dr. Paul Love on the role of the protein Themis in T cell activation. This research usually involves the use of a Zeiss 510 Laser Scanning Confocal Microscope or a PerkinElmer UltraView Spinning Disk Confocal Microscope, however we are seeing increased use of our Total Internal Reflection Fluorescence (TIRF) microscope. Most of the users view immunofluorescent staining on fixed samples with the Zeiss 501 LSCM while the spinning disk confocal is generally used for live cell imaging. We routinely use our TIRF microscope for PhotoActivation Localization Microscopy (PALM). This high resolution technique has allowed us to determine the location of single proteins clustered in signaling complexes in T cells with an accuracy of around 20 nm, well below the diffraction limit of visible light. We can now perform two color PLAM imaging. The TIRF system is also being used for the projects "Regulation of focal adhesions" and "Turnover of invadopodia" with Dr. Paul Randazzo.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Scientific Cores Intramural Research (ZIC)
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Zvezdova, Ekaterina; Lee, Jan; El-Khoury, Dalal et al. (2014) In vivo functional mapping of the conserved protein domains within murine Themis1. Immunol Cell Biol 92:721-8
Coussens, Nathan P; Hayashi, Ryo; Brown, Patrick H et al. (2013) Multipoint binding of the SLP-76 SH2 domain to ADAP is critical for oligomerization of SLP-76 signaling complexes in stimulated T cells. Mol Cell Biol 33:4140-51
Sherman, Eilon; Barr, Valarie A; Samelson, Lawrence E (2013) Resolving multi-molecular protein interactions by photoactivated localization microscopy. Methods 59:261-9
Sherman, Eilon; Barr, Valarie; Samelson, Lawrence E (2013) Super-resolution characterization of TCR-dependent signaling clusters. Immunol Rev 251:21-35
Balagopalan, Lakshmi; Barr, Valarie A; Kortum, Robert L et al. (2013) Cutting edge: cell surface linker for activation of T cells is recruited to microclusters and is active in signaling. J Immunol 190:3849-53
Sommers, Connie L; Gurson, Jordan M; Surana, Rishi et al. (2008) Bam32: a novel mediator of Erk activation in T cells. Int Immunol 20:811-8
Barr, Valarie A; Bernot, Kelsie M; Srikanth, Sonal et al. (2008) Dynamic movement of the calcium sensor STIM1 and the calcium channel Orai1 in activated T-cells: puncta and distal caps. Mol Biol Cell 19:2802-17
Kriebel, Paul W; Barr, Valarie A; Rericha, Erin C et al. (2008) Collective cell migration requires vesicular trafficking for chemoattractant delivery at the trailing edge. J Cell Biol 183:949-61