The following FCB dyes were used: CBD500 (BD Biosciences, San Jose, CA, USA); Pacific Orange NHS ester, DyLight 350 NHS ester, and DyLight 800 NHS ester (Thermo Fisher Scientific, Waltham, MA, USA). Antibodies tested for surface staining were: CD3-BV605 (OKT3) (BioLegend, San Diego, CA); CD4-APC (RPA-T4) (BD Biosciences, San Jose, CA, USA); CD8-PE-Cy5 (B9.11), and tube B of IOTest Beta Mark, containing Vβ 9-PE, Vβ 17-PE/FITC, and Vβ 16-FITC (FIN9, E17.5F3, and TAMAYA1.2) (Beckman Coulter, Miami, FL). LIVE/DEAD Fixable Aqua (a viability dye) for 405 nm excitation was used to exclude dead cells from analysis (Thermo Fisher Scientific). Aqua dye was dissolved in DMSO and stored at −80°C, according to the manufacturer’s instructions. Just before use, Aqua dye was diluted 1:16 with PBS and used for staining. All buffers (Phosflow Lyse/Fix Buffer 5X, Phosflow Perm Buffer II, and Phosflow Barcoding Wash Buffer 4X; BD Biosciences) were prepared, according to the manufacturer’s instructions.
Cd3 bv605 okt3
Manufactured by BioLegend
Sourced in United States
CD3-BV605 (OKT3) is a fluorescently-labeled antibody that binds to the CD3 antigen on the surface of T cells. It can be used for the identification and enumeration of T cells in flow cytometry applications.
Automatically generated - may contain errors
Lab products found in correlation
Cbd500, by BD (1 mentions)
Pacific orange nhs ester, by Thermo Fisher Scientific (1 mentions)
Dylight 350 nhs ester, by Thermo Fisher Scientific (1 mentions)
Cd4 apc rpa t4, by BD (1 mentions)
Live dead fixable aqua, by Thermo Fisher Scientific (1 mentions)
Phosflow lyse fix buffer 5x, by BD (1 mentions)
Cd8 pe cy5, by Beckman Coulter (1 mentions)
Phosflow perm buffer 2, by BD (1 mentions)
Cd19 v500 hib19, by BD (1 mentions)
Iotest beta mark kit, by Beckman Coulter (1 mentions)
Cd56 pe cy7 hcd56, by BioLegend (1 mentions)
Cd16 fitc 3g8, by BD (1 mentions)
Cxcr3 pe, by BioLegend (1 mentions)
2 protocols using cd3 bv605 okt3
1
Multiparametric Flow Cytometry Panel
2
Comprehensive Immune Profiling by Flow Cytometry
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We used flow cytometry with hierarchical gating strategies adapted from established guidelines for analysis of human PBMCs and suitable for cryopreserved samples (18 (link), 19 (link)). Our panel offered survey phenotyping of T cell subsets, including Tregs, as well as B cells and NK cells (see Figure S1 in Supplementary Material). The following fluorochrome-conjugated monoclonal antibodies were used: CD3 BV605 (OKT3, Biolegend), CD4 Alexa 700 (RPA-T4, Biolegend), CD8α V500 (SK1, BD Biosciences), CCR6 PE-Cy7 (G034E3, Biolegend), CXCR3 PE (G025H7, Biolegend), CD127 APC (A019D5, Biolegend), CD25 BV421 (M-A251, Biolegend), CD19 V500 (HIB19, BD Biosciences), CD56 PE-Cy7 (HCD56, Biolegend), CD20 PE (2H7, BD Biosciences), CD14 V450 (MFP9, BD Biosciences), and CD16 FITC (3G8, BD Biosciences). We also used the commercially available IOTest® Beta Mark Kit (Beckman Coulter) for quantitative determination of the human TCR Vβ repertoire in CD4+ and CD8+ cells [nomenclature from Ref. (20 (link))].
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