Researchers at UCLA have developed a fusion protein that can detect immune cells expressing anti-CD19 chimeric antigen receptors with higher specificity and lower background than existing antibodies.
CD19, a glycoprotein specific to the B-cell lineage, is a target of a number of cancer immunotherapies. Clinical trials testing the effectiveness of T cells equipped with anti-CD19 chimeric antigen receptors (CARs) in B-cell malignancy treatments are currently underway. Improving the detection of T cells that carry anti-CD19 CARs would greatly improve the characterization of the cells used for these immunotherapies and increase therapeutic potential.
The detection of CD19 CAR expression by FACS is typically determined through the use of an antibody to either the hinge region or FC portion of the receptor. Detection using commercially available antibodies can result in high levels of background and non-specific binding, especially in blood samples where many lgG expressing cells are present.
Dr. Donald Kohn and colleagues have developed a CD19 fusion protein that can be used to detect anti-CD19 CAR+ cells. This fusion protein can be secreted from producer cells (e.g. 293T) and purified from the cell culture supernatant. The purified protein molecule is conjugated with a fluorescent fluorophore and used directly to detect anti-CD19 CAR+ cells in fluorescence-based techniques, such as flow cytometry (also known as fluorescent activated cell sorting, or FACS) or confocal imaging. This fusion protein contains specific domains of CD19 that should allow the detection of all anti-CD19 CAR+ cells irrespective of the antibody binding domain used to construct the CAR. Immune cells with the anti-CD19 CAR will recognize and specifically bind the CD19 fusion protein, positively labelling the cells. The CD19 domain incorporated into the fusion protein binds anti-CD19 CAR in a highly specific manner and enhances the detection of anti-CD19 CAR+ cells.
fusion protein, antibody, CD19, CAR, flow cytometry, confocal imaging, microscopy, cancer, immunotherapy, oncology, T cell, B cell