Apc anti human cd73 clone ad2

KO cell lines were generated by CRISPR/Cas9-mediated genome editing. A CD47-KO variant of 3LL ΔNRAS cells was generated using Gene Knockout Kit version 2 targeting murine CD47 (Synthego). Knockout variants of PC9, NCI-H358, MGH119, and/or MGH134 were generated using Gene Knockout Kit version 2 targeting human B2M or human CD73 (Synthego). Gene KO was performed via ribonucleoprotein transfection with recombinant Cas9 (Synthego). Cells were then stained for surface antigen expression and sorted using a FACSAria II (BD Biosciences) to generate negative cell lines. For murine CD47, staining was performed using APC anti-murine CD47 clone miap301 (BioLegend) and sorting was used to generate a clonal population. For human lines, staining was performed with APC anti-human B2M clone 2M2 (Biolegend) or APC anti-human CD73 clone AD2 (Biolegend) and used to sort polyclonal lines that were negative for surface antigen expression.

Knockout cell lines were generated by CRISPR/Cas9-mediated genome editing. A CD47 knockout variant of 3LL ΔNRAS cells was generated using Gene Knockout Kit version 2 targeting murine CD47 (Synthego). Knockout variants of PC9, NCI-H358, MGH119, and/or MGH134 were generated using Gene Knockout Kit version 2 targeting human B2M or human CD73 (Synthego). Gene knockout was performed via ribonucleoprotein transfection with recombinant Cas9 (Synthego). Cells were then stained for surface antigen expression and sorted using a FACSAria II (BD Biosciences) to generate negative cell lines. For murine CD47, staining was performed using APC anti-murine CD47 clone miap301 (BioLegend) and sorting was used to generate a clonal population. For human lines, staining wass performed with APC anti-human B2M clone 2M2 (Biolegend) or APC anti-human CD73 clone AD2 (Biolegend) and used to sort polyclonal lines that were negative for surface antigen expression.