siglec 2 fc chimera protein  (R&D Systems)

Journal: Cytotherapy

Article Title: Digital polymerase chain reaction strategies for accurate and precise detection of vector copy number in CAR T cell products

doi: 10.1016/j.jcyt.2022.09.004

Figure Lengend Snippet: Assay #2 was designed at the site between the two distinct CARs within the ORF of the vector, spanning the junction between the linker domains and the 5’-end of the FMC63 scFv of CD19. This primer-probe set was designed to be specific for the bicistronic CD19x22 CAR T cell product. A representative example of droplet readout is pictured here demonstrating that ddPCR confirms specificity of this assay to detect only the bicistronic CAR product as demonstrated with CAR positive droplets (FAM) in blue in the last column. Albumin control (VIC), depicted in green, appropriately demonstrates signal in all samples containing T cells. No CAR positive droplets (in blue) are detected in untransduced T cells or T cells transduced with either CD19 only CARs or CD22 only CARs. NTC = No template control, negative control.

Article Snippet: Following staining and subsequent wash, cells were stained with Recombinant Human CD19-Fc Chimera Atto 647N Protein (R&D Biotechne, ATM9269-020, 1ug/test), Recombinant Human Singlec2/CD22-Fc Chimera Atto 488 (R&D Biotechne, ATJ1968-050, 0.5ug/test), and lyophilized T cell cocktail 6-color Kit (Biolegend, San Diego, CA) containing CD45-PE/Dazzle 594 (clone HI30), CD3 BV510 (clone UCHT1), CD4 Alexa Fluor 700 (clone SK3), CD8 Pacific Blue (clone SK1), CD45RA PE (clone KI100), and CD196 (CCR7) PE/Cy7 (clone G043H7) for 15 minutes at room temperature.

Techniques: Plasmid Preparation, Transduction, Negative Control