We pre-incubated 1 × 106–3 × 106 cells with APC-conjugated VY9, or NP8 tetramer at 37°C, 5% CO2 for one hour. (The tetramers were produced by the NIH Tetramer Core Facility at Emory University, Atlanta, GA.) Next, we added the antibodies recognizing the surface phenotypic markers. We used antibodies from BD Biosciences: CCR7 (clone 150503, R&D Systems) FITC, CD3 (clone SP34-2) PE-CF594, CD8 (clone RPA-T8) BV711, CD20 (clone 2H7) Alexa700, CD28 (clone CD28.2,) PE, CD45 (clone D058-1283) BV786; from BioLegend: PD-1 (clone EH12.2H7) BV605; from Beckman Coulter: NKG2a (clone Z199) PE-Cy7, and Near-Infrared Live/Dead Discriminator from Life Technologies. We incubated the cells for 15 min at room temperature, removed any unbound reagents with two washing steps then fixed them with 2% PFA. We acquired the data on a special-order BD LSR II (BD Biosciences, San Jose, CA) equipped with a 50 mW 405 nm violet, a 100 mW 488 nm blue, and a 50 mW 640 nm red laser, 16 detectors, and FACSDiva software version 8.0.1. We collected up to 250,000 events in the lymphocyte gate defined by the forward and side scatter parameters. We analyzed the data using FlowJo version 10.4.2.
Nkg2a clone z199 pe cy7
Manufactured by Thermo Fisher Scientific
NKG2a (clone Z199) PE-Cy7 is a flow cytometry reagent that binds to the NKG2A receptor on the surface of natural killer (NK) cells and a subset of T cells. It is conjugated with the fluorescent dye PE-Cy7, allowing for the identification and analysis of NKG2A-positive cell populations.
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Lab products found in correlation
Ccr7 clone 150503, by R&D Systems (2 mentions)
Cd3 clone sp34 2 pe cf594, by BioLegend (2 mentions)
Cd8 bv711 clone rpa t8, by BioLegend (2 mentions)
Cd20 clone 2h7 alexa700, by BioLegend (2 mentions)
Pd 1 clone eh12.2h7 bv605, by Beckman Coulter (2 mentions)
Near infrared live dead discriminator, by Thermo Fisher Scientific (2 mentions)
Fluorescein isothiocyanate (fitc), by R&D Systems (2 mentions)
Lsr 2, by BD (2 mentions)
2 protocols using nkg2a clone z199 pe cy7
1
Multiparameter Flow Cytometry Phenotyping
2
Multiparametric Flow Cytometry of T-cell Subsets
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We pre‐incubated 1 × 106–3 × 106 cells with APC‐conjugated VY9, or NP8 tetramer at 37°C, 5% CO2 for one hour. (The tetramers were produced by the NIH Tetramer Core Facility at Emory University, Atlanta, GA.) Next, we added the antibodies recognizing the surface phenotypic markers. We used antibodies from BD Biosciences: CCR7 (clone 150503, R&D Systems) FITC, CD3 (clone SP34‐2) PE‐CF594, CD8 (clone RPA‐T8) BV711, CD20 (clone 2H7) Alexa700, CD28 (clone CD28.2,) PE, CD45 (clone D058‐1283) BV786; from BioLegend: PD‐1 (clone EH12.2H7) BV605; from Beckman Coulter: NKG2a (clone Z199) PE‐Cy7, and Near‐Infrared Live/Dead Discriminator from Life Technologies. We incubated the cells for 15 min at room temperature, removed any unbound reagents with two washing steps then fixed them with 2% PFA. We acquired the data on a special‐order BD LSR II (BD Biosciences, San Jose, CA) equipped with a 50 mW 405 nm violet, a 100 mW 488 nm blue, and a 50 mW 640 nm red laser, 16 detectors, and FACSDiva software version 8.0.1. We collected up to 250,000 events in the lymphocyte gate defined by the forward and side scatter parameters. We analyzed the data using FlowJo version 10.4.2.
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