CAR Mesothelin surface expression on human T-cells was assessed using primary antibody goat anti-human IgG F (ab’)2 Biotin (BioRad, Hercules, CA) along with secondary conjugate Streptavidin APC (BioRad). The following antibodies were also used in FACS buffer in this study: anti-PD1-PE (Biolegends 329906), anti-CCR7-Alexa-fluor 647 (BD 560921), anti-CD8-FITC (BC A07756), anti-CD8-BV421 (BD 562428), anti-HLA-A2-FITC (BD 551,285), anti-CD86-PerCp-Cy5.5 (BD 561129), anti-CLDN6-Dylight 650 (IMAB027), anti-idiotype-IMAB027- Alexa-fluor 647, anti-TIM3-APC (Biolegends 345011) and 7AAD (BC A07704). Mesothelin expression was detected using the PE conjugated anti mesothelin antibody (R&D system FAB32652P). Acquisition and analysis of all samples were performed on a BD FACS CantoI/II (BD Biosciences, San Jose, California) and FlowJo software (v7.6.1).
Anti tim3 apc
Manufactured by BioLegend
Anti-TIM3-APC is a monoclonal antibody that binds to the T-cell immunoglobulin and mucin domain-containing protein 3 (TIM3). TIM3 is an inhibitory receptor expressed on T cells, natural killer cells, and dendritic cells that plays a role in regulating immune responses.
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Lab products found in correlation
Anti pd 1 pe, by BioLegend (2 mentions)
Anti cd8 bv421, by BD (1 mentions)
Goat anti human igg f ab 2 biotin, by Bio-Rad (1 mentions)
Streptavidin apc, by Bio-Rad (1 mentions)
Anti cd86 percp cy5, by BD (1 mentions)
Facs cantoi 2, by BD (1 mentions)
Anti hla a2 fitc, by BD (1 mentions)
Anti ccr7 alexa fluor 647, by BD (1 mentions)
Fab32652p, by R&D Systems (1 mentions)
Facsdiva software, by BD (1 mentions)
Anti cd44, by Thermo Fisher Scientific (1 mentions)
Anti cd24, by Thermo Fisher Scientific (1 mentions)
Anti cd69, by Thermo Fisher Scientific (1 mentions)
Fortessa flow cytometer, by BD (1 mentions)
Anti foxp3 alexa 488, by BioLegend (1 mentions)
Anti cd8, by BioXCell (1 mentions)
Anti cd107a pe cy7, by BioLegend (1 mentions)
Anti ifn γ bv650, by BioLegend (1 mentions)
Anti f4 80 pecy7, by BioLegend (1 mentions)
Anti cd86 bv605, by BioLegend (1 mentions)
3 protocols using anti tim3 apc
1
Mesothelin Expression on Human T-cells
2
Multiparameter Flow Cytometry Protocol
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The following monoclonal Abs were used in flow cytometry: anti-CD45 (30–11, APC-Cy7, PE), anti-CD8 (Ly-2, V450), anti-CD4 (RM4-5, V500) anti-CD44 (IM7, Alexa 700) anti-mouse OVA257-264 (SIINFEKL) peptide bound to H-2Kb (PE), anti-CD326 (EPCAM) (9C4; FITC), anti-CD24 (30-F1, PE-Cy7), anti-CD69 (H1.2F3, PerCP/Cy5.5), anti-CD61 (Ebioscience, 2C9.G3; FITC) anti-PDGRFα CD140α (Abcam, 16A1; PE); anti-Ly6C PE, anti-Ly6G BV785, anti-Foxp3 Alexa 488, anti-F4/80 Pe-Cy7, anti-CD206 BV650, anti-CD11b Percp Cy5, anti-CD86 BV605, anti-CD80 APC-Cy7, anti-IFNγBV650, anti-GrzB Alexa647, anti-TNFα BV605, anti-NKp46 Alexa 488, anti-CD107a Pe-Cy7, anti-CD29 Percp Cy5, anti-PD-1 PE, anti-Tim3 APC, anti-Lag3 Percp Cy5 and anti-CD11c Alexa 700 (all from Biolegend). The flow cytometry analyses were performed using a BD Fortessa Flow Cytometer (BD Biosciences) and analysed using either BD FACS Diva software V.5.0.1 (BD) or FlowJo (Tree Star). For the in vivo studies, the following endotoxin-free Abs were used: anti-CD8 (BioXcell; 2.43), anti-βigh3 18B3,51 (link) anti-PD-1 and control polyclonal mouse Ig (BioXcell).
3
Comprehensive Flow Cytometry Immunophenotyping
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For flow cytometry analysis, tumor and blood samples were stained with a cocktail of monoclonal mouse anti-human conjugated antibodies (mAbs): anti-CD45-PercP (clone HI30), anti-CD3-PercP (HIT3a), anti-CD3-APC (UCHT1), anti-CD19-PE (HIB19), anti-CD15-PE (HI98), anti-CD161-FITC (HP-3G10), anti-CD4-FITC (OKT4), anti-CD8-PE (HIT8a), anti-HLA-DR-APC (L243), anti-CD127-PE-Cy7 (A019D5), anti-CD1c-APC-Cy7 (L161), anti-CD163-PE (GHI/61), anti-CD206-APC-Cy7 (15–2), anti-PD-1-FITC (EH12.2H7), anti-PD-L1-APC (29E2A3), anti-CTLA4-PE (L3D10), anti-CD69-APC-Cy7 (FN50), anti-Tim3-APC (F38-2E2), anti-IL-8-APC (E8N1), anti-IFN-γ-PE (4S.B3), anti-IFN-γ-APC-Cy7 (4S.B3), anti-Granzyme B-FITC (QA16A02), anti-IL-1β-FITC (JK1B-1), anti-IL-2-PE-Cy7 (MQ1-17H12), anti-IL-6-APC (MQ2-13A5), anti-IL-17-FITC (BL168), anti-IL-23/IL-12-PE (C11.5), anti-TGF-β-APC (TW4-6H10), all from Biolegend; anti-IDO-PE (eyedio) and anti-IL-10-FITC (BT-10), both from eBioscience; anti-CD25-PE (MEM-181) and anti-CD11b-FITC (LT11) from ImmunoTools.
The antibodies used for immunofluorescence were: mouse monoclonal anti-human CD8 (32-M4) from Santa Cruz Biotechnology and rabbit polyclonal anti-human HLA-DRA from Sigma Aldrich.
The antibodies used for immunofluorescence were: mouse monoclonal anti-human CD8 (32-M4) from Santa Cruz Biotechnology and rabbit polyclonal anti-human HLA-DRA from Sigma Aldrich.
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