Flow Cytometry Unit
Flow cytometry is a powerful analysis tool in modern biology and life sciences. The applications are numerous and have contributed to many fields including immunology, cell biology and microbiology. The flow cytometer performs the analysis by passing thousands of cells per second through laser beams and by capturing the light that emerges from each cell as it passes through. The gathered data can be analyzed statistically by flow cytometry software to report cellular characteristics such as size, complexity, phenotype, and health. In addition, defined subpopulations of cells can be purified for further experiments according to their phenotype.
The Flow Cytometry Unit provides cell analysis and sorting services to all departments and groups within the Institute. Available techniques include high-speed cell sorting, multi-color immunostaining, expression analysis of fluorescent proteins, DNA ploidy and cell cycle analysis, single cell cloning as well as measurement of apoptosis and fluorescence resonance energy transfer (FRET). Sterile sorting allows subsequent culturing of sorted cells. The service further includes assistance with analysis, interpretation and professional presentation of flow cytometric data.
The Flow Cytometry Unit operates two high-speed cell sorters: a FACSAria IIIu cell sorter (BD Biosciences) equipped with five lasers (375 nm, 405 nm, 488 nm, 561 nm and 633 nm) and 13 fluorescence detectors, and a FACSAria Fusion with five lasers (355 nm, 405 nm, 488 nm, 561 nm and 633 nm) and 18 fluorescence detectors. Both instruments are capable of detecting almost any commercially available fluorescent cell labels. Up to four subpopulations can be sorted out in parallel. An Automatic Cell Deposition Unit (ACDU) allows sorting of single cells into multi-well plates.
Training and supervision is provided to researchers who wish to operate our cell sorters on their own. Furthermore license dongles for flow cytometry analysis software (FlowJo, Kaluza) can be provided.
Publications
Chen Q, Liu Y, Jeong HW, Stehling M, Dinh VV, Zhou B, Adams RH (2019) Apelin+ Endothelial Niche Cells Control Hematopoiesis and Mediate Vascular Regeneration after Myeloablative Injury. Cell Stem Cell 25(6):768-783
Skvortsova K, Tarbashevich K, Stehling M, Lister R, Irimia M, Raz E, Bogdanovic O (2019) Retention of paternal DNA methylome in the developing zebrafish germline. Nat Commun 10(1):3054
Reinhardt L, Kordes S, Reinhardt P, Glatza M, Baumann M, Drexler HCA, Menninger S, Zischinsky G, Eickhoff J, Fröb C, Bhattarai P, Arulmozhivarman G, Marrone L, Janosch A, Adachi K, Stehling M, Anderson EN, Abo-Rady M, Bickle M, Pandey UB, Reimer MM, Kizil C, Schöler HR, Nussbaumer P, Klebl B, Sterneckert JL (2019) Dual Inhibition of GSK3β and CDK5 Protects the Cytoskeleton of Neurons from Neuroinflammatory-Mediated Degeneration In Vitro and In Vivo. Stem Cell Reports 12(3):502-517
Cavaco ACM, Rezaei M, Caliandro MF, Lima AM, Stehling M, Dhayat SA, Haier J, Brakebusch C, Eble JA (2018) The Interaction between Laminin-332 and α3β1 Integrin Determines Differentiation and Maintenance of CAFs, and Supports Invasion of Pancreatic Duct Adenocarcinoma Cells. Cancers 11(14)
Adachi K, Kopp W, Wu G, Heising S, Greber B, Stehling M, Araúzo-Bravo MJ, Boerno ST, Timmermann B, Vingron M, Schöler HR (2018) Esrrb Unlocks Silenced Enhancers for Reprogramming to Naive Pluripotency. Cell Stem Cell 23(6):900-904
Hägerling R, Hoppe E, Dierkes C, Stehling M, Makinen T, Butz S, Vestweber D, Kiefer F (2018) Distinct roles of VE-cadherin for development and maintenance of specific lymph vessel beds. EMBO J 15;37(22)
Kato K, Diéguez-Hurtado R, Park DY, Hong SP, Kato-Azuma S, Adams S, Stehling M, Trappmann B, Wrana JL, Koh GY, Adams RH (2018) Pulmonary pericytes regulate lung morphogenesis. Nat Commun 9(1):2448
Weider M, Starost LJ, Groll K, Küspert M, Sock E, Wedel M, Fröb F, Schmitt C, Baroti T, Hartwig AC, Hillgärtner S, Piefke S, Fadler T, Ehrlich M, Ehlert C, Stehling M, Albrecht S, Jabali A, Schöler HR, Winkler J, Kuhlmann T, Wegner M (2018) Nfat/calcineurin signaling promotes oligodendrocyte differentiation and myelination by transcription factor network tuning. Nat Commun 9(1):899
Gross-Thebing T, Yigit S, Pfeiffer J, Reichman-Fried M, Bandemer J, Ruckert C, Rathmer C, Goudarzi M, Stehling M, Tarbashevich K, Seggewiss J, Raz E (2017) The Vertebrate Protein Dead End Maintains Primordial Germ Cell Fate by Inhibiting Somatic Differentiation. Dev Cell 43(6):704-715
Pitulescu ME, Schmidt I, Giaimo BD, Antoine T, Berkenfeld F, Ferrante F, Park H, Ehling M, Biljes D, Rocha SF, Langen UH, Stehling M, Nagasawa T, Ferrara N, Borggrefe T, Adams RH (2017) Dll4 and Notch signalling couples sprouting angiogenesis and artery formation. Nat Cell Biol 19(8):915-927
Seebeck F, März M, Meyer AW, Reuter H, Vogg MC, Stehling M, Mildner K, Zeuschner D, Rabert F, Bartscherer K (2017) Integrins are required for tissue organization and restriction of neurogenesis in regenerating planarians Development 144(5):795-807
Chen Q, Zhang H, Liu Y, Adams S, Eilken H, Stehling M, Corada M, Dejana E, Zhou B, Adams RH (2016) Endothelial cells are progenitors of cardiac pericytes and vascular smooth muscle cells. Nat Commun 7:12422
Konigsberger S, Weis V, Prodohl J, Stehling M, Hobeika E, Reth M, Kiefer F (2015). Suboptimal B-cell antigen receptor signaling activity in vivo elicits germinal center counterselection mechanisms. Eur J Immunol 45(2): 603-611
Sugawa F, Arauzo-Bravo MJ, Yoon J, Kim KP, Aramaki S, Wu G, Stehling M, Psathaki OE, Hubner K, Scholer HR (2015). Human primordial germ cell commitment in vitro associates with a unique PRDM14 expression profile. EMBO J 34(8): 1009-1024
Volkenhoff A, Weiler A, Letzel M, Stehling M, Klambt C, Schirmeier S (2015). Glial Glycolysis Is Essential for Neuronal Survival in Drosophila. Cell Metab 22(3): 437-447
Baumer N, Baumer S, Berkenfeld F, Stehling M, Kohler G, Berdel W E, Muller-Tidow C, Tschanter P (2014). Maintenance of leukemia-initiating cells is regulated by the CDK inhibitor Inca1. PLoS One 9(12): e115578
Hartwig J, Tarbashevich K, Seggewiss J, Stehling M, Bandemer J, Grimaldi C, Paksa A, Gross-Thebing T, Meyen D, Raz E (2014). Temporal control over the initiation of cell motility by a regulator of G-protein signaling. Proc Natl Acad Sci U S A 111(31): 11389-11394
Joo JY, Choi HW, Kim MJ, Zaehres H, Tapia N, Stehling M, Jung KS, Do JT, Scholer HR (2014). Establishment of a primed pluripotent epiblast stem cell in FGF4-based conditions. Sci Rep 4: 7477
Kim J, Eligehausen S, Stehling M, Nikol S, Ko K, Waltenberger J, Klocke R (2014). Generation of functional endothelial-like cells from adult mouse germline-derived pluripotent stem cells. Biochem Biophys Res Commun 443(2): 700-705
Singhal N, Esch D, Stehling M, Scholer HR (2014). BRG1 Is Required to Maintain Pluripotency of Murine Embryonic Stem Cells. Biores Open Access 3(1): 1-8
Pfeiffer MJ, Esteves TC, Balbach ST, Arauzo-Bravo MJ, Stehling M, Jauch A, Houghton FD, Schwarzer C, Boiani M (2013). Reprogramming of two somatic nuclei in the same ooplasm leads to pluripotent embryonic stem cells. Stem Cells 31(11): 2343-2353
Thyrock A, Ossendorf E, Stehling M, Kail M, Kurtz T, Pohlentz G, Waschbusch D, Eggert S, Formstecher E, Muthing J, Dreisewerd K, Kins S, Goud B, Barnekow A (2013). A new Mint1 isoform, but not the conventional Mint1, interacts with the small GTPase Rab6. PLoS One 8(5): e64149
Wang L, Benedito R, Bixel M G, Zeuschner D, Stehling M, Savendahl L, Haigh JJ, Snippert H, Clevers H, Breier G, Kiefer F, Adams RH (2013). Identification of a clonally expanding haematopoietic compartment in bone marrow. EMBO J 32(2): 219-230
Höing S, Rudhard Y, Reinhardt P, Glatza M, Stehling M, Wu G, Peiker C, Böcker A, Parga JA, Bunk E, Schwamborn JC, Slack M, Sterneckert J, Schöler HR (2011). Discovery of inhibitors of microglial neurotoxicity acting through multiple mechanisms using a stem-cell-based phenotypic assay Cell Stem Cell 11(5):620-32
Königsberger S, Prodöhl J, Stegner D, Weis V, Andreas M, Stehling M, Schumacher T, Böhmer R, Thielmann I, van Eeuwijk JM, Nieswandt B, Kiefer F (2011). Altered BCR signalling quality predisposes to autoimmune disease and a pre-diabetic state EMBO J 31(15):3363-74
Elling C, Erben P, Walz C, Frickenhaus M, Schemionek M, Stehling M, Serve H, Cross NC, Hochhaus A, Hofmann WK, Berdel WE, Müller-Tidow C, Reiter A, Koschmieder S (2011). Novel imatinib-sensitive PDGFRA-activating point mutations in hypereosinophilic syndrome induce growth factor independence and leukemia-like disease Blood 117(10):2935-43
Han DW, Greber B, Wu G, Tapia N, Araúzo-Bravo MJ, Ko K, Bernemann C, Stehling M, Schöler HR (2011). Direct reprogramming of fibroblasts into epiblast stem cells. Nat Cell Biol 13(1):66-71
Sabour D, Araúzo-Bravo MJ, Hübner K, Ko K, Greber B, Gentile L, Stehling M, Schöler HR (2011). Identification of genes specific to mouse primordial germ cells through dynamic global gene expression. Hum Mol Genet 1;20(1):115-25
Steffen B, Knop M, Bergholz U, Vakhrusheva O, Rode M, Köhler G, Henrichs MP, Bulk E, Hehn S, Stehling M, Dugas M, Bäumer N, Tschanter P, Brandts C, Koschmieder S, Berdel WE, Serve H, Stocking C, Müller-Tidow C (2011). AML1/ETO induces self-renewal in hematopoietic progenitor cells via the Groucho-related amino-terminal AES protein Blood 117(16):4328-37
Agrawal-Singh S, Koschmieder S, Gelsing S, Stocking C, Stehling M, Thiede C, Thoennissen NH, Köhler G, Valk PJ, Delwel R, Mills K, Bäumer N, Tickenbrock L, Hansen K, Berdel WE, Müller-Tidow C, Serve H (2010). Pim2 cooperates with PML-RARalpha to induce acute myeloid leukemia in a bone marrow transplantation model. Blood 115(22):4507-16
Böhmer R, Neuhaus B, Bühren S, Zhang D, Stehling M, Böck B, Kiefer F (2010). Regulation of developmental lymphangiogenesis by Syk(+) leukocytes. Dev Cell 18(3):437-49
Fernandéz-Taboada E, Moritz S, Zeuschner D, Stehling M, Schöler HR, Saló E, Gentile L (2010). Smed-SmB, a member of the LSm protein superfamily, is essential for chromatoid body organization and planarian stem cell proliferation. Development 137(7):1055-65
Han DW, Tapia N, Joo JY, Greber B, Araúzo-Bravo MJ, Bernemann C, Ko K, Wu G, Stehling M, Do JT, Schöler HR (2010). Epiblast stem cell subpopulations represent mouse embryos of distinct pregastrulation stages. Cell 143(4):617-27
Ko K, Araúzo-Bravo MJ, Kim J, Stehling M, Schöler HR (2010). Conversion of adult mouse unipotent germline stem cells into pluripotent stem cells. Nat Protoc 5(5):921-8
Schemionek M, Elling C, Steidl U, Bäumer N, Hamilton A, Spieker T, Göthert JR, Stehling M, Wagers A, Huettner CS, Tenen DG, Tickenbrock L, Berdel WE, Serve H, Holyoake TL, Müller-Tidow C, Koschmieder S (2010). BCR-ABL enhances differentiation of long-term repopulating hematopoietic stem cells. Blood 115(16):3185-95
Sterneckert J, Stehling M, Bernemann C, Araúzo-Bravo MJ, Greber B, Gentile L, Ortmeier C, Sinn M, Wu G, Ruau D, Zenke M, Brintrup R, Klein DC, Ko K, Schöler HR (2010). Neural induction intermediates exhibit distinct roles of Fgf signaling. Stem Cells 28(10):1772-81
Thyrock A, Stehling M, Waschbüsch D, Barnekow A (2010). Characterizing the interaction between the Rab6 GTPase and Mint3 via flow cytometry based FRET analysis. Biochem Biophys Res Commun 396(3):679-83
Ko K, Tapia N, Wu G, Kim JB, Bravo MJ, Sasse P, Glaser T, Ruau D, Han DW, Greber B, Hausdörfer K, Sebastiano V, Stehling M, Fleischmann BK, Brüstle O, Zenke M, Schöler HR (2009) Induction of pluripotency in adult unipotent germline stem cells. Cell Stem Cell 2;5(1):87-96
Zucchi I, Astigiano S, Bertalot G, Sanzone S, Cocola C, Pelucchi P, Bertoli G, Stehling M, Barbieri O, Albertini A, Schöler HR, Neel BG, Reinbold RA, Dulbecco R (2008) Distinct populations of tumor-initiating cells derived from a tumor generated by rat mammary cancer stem cells. Proc Natl Acad Sci 4;105(44):16940-5
Do JT, Han DW, Gentile L, Sobek-Klocke I, Stehling M, Schöler HR (2008) Enhanced reprogramming of Xist by induced upregulation of Tsix and Dnmt3a. Stem Cells 26(11):2821-31
Cantz T, Bleidissel M, Stehling M, Schöler HR (2008) In vitro differentiation of reprogrammed murine somatic cells into hepatic precursor cells. Biol Chem. 389(7):889-96
Han DW, Do JT, Gentile L, Stehling M, Lee HT, Schöler HR. (2008) Pluripotential reprogramming of the somatic genome in hybrid cells occurs with the first cell cycle. Stem Cells 26(2):445-54
Rustemeyer P, Wittkowski W, Greve B, Stehling M (2007) Flow-cytometric identification, enumeration, purification, and expansion of CD133+ and VEGF-R2+ endothelial progenitor cells from peripheral blood. J Immunoassay Immunochem. 28(1):13-23
Do JT, Han DW, Gentile L, Sobek-Klocke I, Stehling M, Lee HT, Schöler HR (2007) Erasure of cellular memory by fusion with pluripotent cells. Stem Cells 25(4):1013-20