Pre-made Lentivirus express the CAR construct of “anti-CD19-ScFV (VL-VH)-CD8 hinge-CD28-CD3ζ”, containing RFP–Puromycin (GFP-Puro), Fluorescent-Antibiotic dual selection that allow to select the positive trnasduced cells via RFP signal or Puromycin killing.
The ScFv of anti-CD19 (clone FMC63) fusion with the CAR (Chimeric Antigen Receptor) of “CD8 hinge, 4-1BB co-stimulatory domain, and CD3ζ signaling domains”, is expressed under the enhance EF1a promoter. The lentivector’s core structure is represented by the scheme below. It is used as CAR-T targeted killing of the CD19 positive cancer cells. see details in Product Manual.
CD19 is a marker of in most B cell leukemias and lymphomas but not in any normal tissue other than the B cell lineage, CD19 is used to diagnose cancers that arise from this type of cell, and is used as the target for CD19-targeted therapies.
CD28 is transmembrane protein expressed on T cells. It provides co-stimulatory signals for inducing T cell activation and proliferation. CD28 costimulatory domains in CARs led to enhanced anti-malignancy efficacy. CD28 is widely used as the costimulatory domain in CARs.
CD3ζ (T-cell receptor zeta) is expressed by T cells and NK cells. It together with T-cell receptor and CD3γ, δ, ε chain, forms the TCR-CD3 complex. CD3-zeta is the most commonly used activation component of CARs. It transmits an activation signal to the T cell after the antigen is bound. It can be coupled with additional co-stimulatory signaling for the complete activation.
Negative Controls: To validate the specificity of the targeted CAR-T killing, you can use the two negative controls CAR lentivirus: the CAR-ctr3 that has the Anti-CD-19-ScFv recognization removed, or the CAR-Ctr4 that has the CD3ζ activation domain removed, from the CAR construct with puromycin selection.
Amount: 200 ul/vial at titer of 1×108 IFU/ml, concentrated lentivirus provided in PBS solution, premixed with 10x polybrene (60ug/ml), premade, ready to ship, in dry-ice package.