Planar bilayers were constructed as above described and incubated with primary human CD3+ CD4+ CD27+ CD28+ T cells. After 20 min activation on bilayers, CD3+ CD4+ CD27+ CD28+ T cells were removed with cold PBS. The released synaptic TCRs were retained on the bilayer, as described19 (link). Next, telomere-live-labelled APCs (obtained by introducing and TelC probe by glass beads as above described) were loaded with antigen pool and transferred on the TCR coated planar bilayers. The % of APCs releasing telomeres on bilayers was quantified 24h later. In control experiments, APCs were not loaded with antigen pool; or they were transferred onto bilayers constructed with ICAM-1 molecules alone without UCTH1. The vesicular nature of the released telomeres was further tested by pre-incubating TelC labelled APCs with antibodies to the vesicle marker CD63 (clone H5c6; Biolegend Cat. 353015) at 1:50 dilution prior to transfer on bilayer.
Clone h5c6
Manufactured by BioLegend
Sourced in United States
Clone H5c6; is a laboratory reagent used for the detection and analysis of specific cellular markers. It serves as a tool for researchers to identify and study target cells or molecules within a sample.
Automatically generated - may contain errors
Lab products found in correlation
Clone dreg 56, by BioLegend (1 mentions)
Clone fun 2, by BioLegend (1 mentions)
Clone g10f5, by BioLegend (1 mentions)
Clone 12g5, by BioLegend (1 mentions)
Facscelesta, by BD (1 mentions)
Facsdiva software, by BD (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Thermo Fisher Scientific (1 mentions)
3 protocols using clone h5c6
1
Telomere Transfer Dynamics in T-APC Synapses
2
Telomere Transfer Dynamics in T-APC Synapses
3
Phenotypic Profiling of Neutrophils
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Isolated neutrophils (50,000 cells) were stained in fluorescent activated cell sorting (FACS) buffer containing 2% heat-inactivated fetal bovine serum (FBS) (Life Technologies, Dun Laoghaire, Ireland) and 1 mM EDTA (Life Technologies) in phosphate buffer saline (PBS) without calcium and magnesium (Corning, Corning, NY, USA). Cells were stained with antibodies for 30 min at 4 °C in FACS buffer containing (BV421) anti-CD10 (1:200 dilution; clone HI10a; BioLegend, San Diego, CA, USA) and (BV786) anti-CXCR4 (1:100 dilution; clone12G5; BioLegend) and (AF647) anti-CD64 (1:100 dilution; clone 10.1; BioLegend) or (BV786) anti-CD63 (1:100 dilution; clone H5C6; BioLegend) and (APC) anti-CD66b (1:800 dilution; clone G10F5; BioLegend), or (BV421) anti-CD62L (1:200 dilution; clone DREG-56; BioLegend), and (PE/Dazzle) anti-CD32 (1:200 dilution; clone FUN-2; BioLegend). Data were acquired on a BD FACSCelesta (BD Biosciences, San Jose, CA, USA) with a BVR laser configuration (488 nm, 405 nm, 640 nm). Before recording data, gates were prepared so that 10,000 neutrophil events could be collected. FCS files were exported from BD FACSDiva Software (BD Biosciences) in a 3.0 format. FCS files were analyzed using FlowJo v.10 software (BD Biosciences).
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