For surface staining, cells (1–5 × 105) were harvested from culture and washed with FACS buffer (PBS with 1% of BSA and 0.1% sodium azide) prior to staining. The cells were treated with human FcR blocking reagent (Miltenyi Biotec) on ice for 20 minutes, and cells were then treated with various fluorescent conjugated antibodies against cell surface markers in FACS buffer. Human γδ T cells were stained with APC-conjugated anti-human TCRγ/δ (Biolegend, clone: B1) or APC-conjugated anti-human TCRα/β (Biolegend, clone: IP26) and Pacific Blue-conjugated anti-human CD3 (Biolegend, clone: OKT3). For influenza-specific tetramer staining, cells (1–5 × 105) were stained with APC-conjugated Influenza-M1 tetramer (Beckman Coulter, Brea, CA, HLA-A*0201, GILGFVFTL) in FACS buffer at room temperature for 1 hour prior to surface staining with the same staining antibodies described.
Pacific blue conjugated anti human cd3
Manufactured by BioLegend
Pacific Blue-conjugated anti-human CD3 is a fluorochrome-conjugated antibody that binds to the CD3 antigen expressed on human T cells. This product can be used for the identification and enumeration of T cells in flow cytometry applications.
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Lab products found in correlation
Human fcr blocking reagent, by Miltenyi Biotec (1 mentions)
Apc conjugated anti human tcrα β, by BioLegend (1 mentions)
Cyan adp, by Agilent Technologies (1 mentions)
Live dead fixable aqua dead cell stain kit, by Thermo Fisher Scientific (1 mentions)
Cytofix cytoperm buffer, by BD (1 mentions)
Pe cy7 conjugated anti human cd3, by BioLegend (1 mentions)
2 protocols using pacific blue conjugated anti human cd3
1
Cell Surface Staining and Tetramer Assays
2
Multiparameter Flow Cytometry for HIV-1 Infection
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For HIV-1 gag p24 staining, cells were stained with surface markers first, and then permeabilized with cytofix/cytoperm buffer (BD Bioscience, cat#554714), followed by intracellular staining. Human leukocytes (mCD45−huCD45+) were analyzed for human CD3, CD4, CD8, CD123, HLA-DR and CD38 by CyAn FACS machine (Dako). FITC-conjugated anti–human HLA-DR (clone:L243, cat#307604), PE-conjugated anti-human CD38 (clone:HIT2, cat#303506), PE/Cy5-conjugated anti-human CD4 (clone:RP4-T4, cat#300510), PE/Cy7-conjugated anti-human CD3 (clone:HIT3a, cat#300316), Pacific blue-conjugated anti-human CD3 (clone:UCHT1, cat#300431), PE/Cy7-conjugated anti-human CD8 (clone:HIT8a, cat#300914), APC-conjugated human CD123 (clone:6H6, cat#306012) and APC/Cy7-conjuaged anti-human CD45 (clone:H130, cat#304014) were purchased from Biolegend; PE-conjugated anti-human caspase-3 (clone:C92-605, cat#51-68655X) was purchased from BD Bioscience. Pacific orange–conjugated anti–mouse CD45 (clone:HI30, cat#MHCD4530), PE/Texas red–conjugated anti–human CD4 (clone:S3.5, cat#MHCD0417) or CD8 (clone:3B5, cat#MHCD0817), and LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (cat#L34957) were purchased from Invitrogen. FITC-conjugated anti-HIV p24 (clone:FH190-1-1, cat#6604665) was purchased from Beckman Coulter. The cells were analyzed on a CyAn ADP (Dako).
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