Apc anti human cd62l antibody

To assess the influence of EGFRvIII antigen-dependent activation, proliferation, differentiation and exhaustion on T cell phenotypes. Briefly, after coincubation EGFRvIII Nb-CAR-T cells with the target EGFRvIII⁺ cells, T cells surface phenotypes were assessed using the following antibodies: PE or APC anti-human CD25 antibody (BioLegend, USA), PE or APC anti-human CD69 antibody (BioLegend, USA) for T cell activation; PKH26 (Sigma Aldrich, USA) or eFluor670 dye (ThermoFisher, USA) for T cell proliferation; APC anti-human CD62L antibody (BioLegend, USA) and PE-cy7 anti-human CD45RA antibody (BioLegend, USA) for T cell memory function; APC anti-human LAG-3 antibody (BioLegend, USA) and APC anti-human TIM-3 antibody (BioLegend, USA) for T cell exhaustion, all flow cytometry data was analyzed using FlowJo software.

All chemicals are from Sigma-Aldrich (highest grade) if not mentioned otherwise. The following antibodies and reagents were purchased from Biolegend: APC/Cy7 anti-human CD3 antibody, BV421 anti-human CD3 antibody, BV421 anti-human CD8 antibody, APC anti-human CD253 (TRAIL) antibody, APC anti-mouse CD253 (TRAIL) Antibody, BV421 anti-mouse CD8a Antibody, PE anti-mouse CD3 Antibody, PerCP anti-human CD25 antibody, APC anti-human CD62L antibody, APC anti-human CD262 (TRAIL-R2), and 7-AAD viability staining solution. The following antibodies were also used: FITC anti-human CD69 (eBiosciences), FITC anti-human CD44 (DAKO), Purified NA/LE mouse anti-human CD253 (BD Biosciences), BV421 mouse anti-human CD263 (TRAIL-R3) (BD Biosciences), Alexa647 mouse anti-human GLUT1 (BD Biosciences), human TRAIL R1/TNFRSF 10A PerCP-conjugated antibody (R&D Systems), and human TRAIL R4/TNFRSF 10D PE-conjugated antibody (R&D Systems). In addition, the following reagents were used: NucView Caspase-3 enzyme substrates (Biotium), Idelalisib (Selleckchem), MK-2206 (Selleckchem), Rapamycin (Selleckchem), Caffeic acid phenethyl ester (CAPE) (R&D Systems), Mitoquinone (MitoQ) (Biotrend), N-acetyl-L-cysteine (NAC) (Merck), vitamin 1,25D3 (Merck), Calcipotriol (TOCRIS), DMSO (Merck), Cellular ROS Assay Kit (Abcam), metformin hydrochloride (Merck), H₂O₂ (Merck), and Streptozotocin (Merck).