Whole Blood Assay with Dual Co-Stimulation for Antigen-Specific Analysis of Host Immunity to Fungal and Viral Pathogens

Our research focuses on the immune monitoring of patients in the intensive care unit. Some patients develop severe viral pneumonia during mechanical ventilation. We isolate the cause of the viruses, but also characterize the local and systemic immune response using the protocol presented in this video to identify immunological risk factors.

Traditional antigen-specific T-cell assays that are based on isolated mononuclear cells are highly susceptible to pre-analytical confounders, time-and resource-intensive, and require large amounts of blood. This poses severe limitations to the implementation of antigen-specific T-cell assays in the clinical research or clinical routine use as prognostic or diagnostic biomarkers. We present a versatile immunoassay protocol, which accommodates various downstream readouts from as little as 250 micros blood per stimulus.

This enables applications, even when that volume is limited or several antigens need to be screened. Furthermore, our assay is easy to perform and amenable to several pathogens. In addition to its versatility and the low blood volume required, our protocol used as stimulation environment that was thoroughly and specifically optimized for antigen-induced T-helper cell responses.

This was achieved through dual core stimulation, which also makes the protocol more robust towards confounders, such as immunosuppressive pharmacotherapy or pre-analytical delays. We are using this protocol for immune monitoring of patients with ventilation-associated viral pneumonia. Our collaborators in Wuerzburg and Houston are working on a clinical translation of cytomegalovirus-specific immunoassays.

Furthermore, they're using this protocol for studies on fungal immunopathogenesis and preclinical immunotherapy studies in mouse models for opportunistic infections.