The study of cellular binding is needed to identify if a drug binds to the expected cell types or instead binds nonspecifically to any of the blood cells. Our platform provides a unique opportunity to track the binding of different cell populations simultaneously. The binding of an antibody or fluorophore-labeled drug to blood cells can be studied at various time points in a physiologically relevant environment with correct target and Fc-receptor expression present. This allows the investigation of downstream processes related to drug binding, such as cell activation, cargo uptake, internalization, shedding, trogocytosis and phagocytosis. Surface or intracellular location can be identified through quenching of surface staining or the use of pH-sensitive dyes. The binding profile in the circulation over time can be compared with the binding profile noted in the fresh static blood directly after collection in conditions that stop protein traffic, shedding or internalization processes, which can be a valuable complement to PK/PD analysis.
Freshly acquired whole blood was incubated with anti-integrin alpha 4 (2 µg/ml) or vehicle using Immuneed’s platform based on circulating blood. Blood acquired directly after collection (zero time point) and at the 4-hour time point was stained with fluorescently labelled monoclonal antibodies to detect: erythrocytes (CD235ab+, CD45-), platelets (CD41+, CD45-), T cells (CD3+), B cells (CD19+), NK cells (CD56+), monocytes (CD14+) and granulocytes (CD66b+).