Integrated platforms for automatic assessment of cellular functional secretory immunophenotyping could have a widespread use in the diagnosis, real-time monitoring, and therapy evaluation of several pathologies. We present a microfluidic platform with integrated biosensors and culture chambers to measure cytokine secretion from a consistent and uniform number of immune cells. The biosensor relies on a fluorescence sandwich immunoassay enabled by the mechanically induced trapping of molecular interactions method. The platform contains 32 cell culture chambers, each patterned with an array of 492 microwells, to capture and analyze both adherent and nonadherent immune cells. Multiple stimuli can be delivered to a set of culture chambers. Per chamber, we were able to capture consistently 1113 ± 191 of blood-derived monocytes and neutrophils and 348 ± 37 THP-1 monocytes. Good occupancy efficiencies of ∼70% with a uniformity of ∼90% across all of the culture chambers of the device were achieved. Furthermore, we demonstrate that up to 96% of cells remain viable for the first 48 h. The employment of epoxy-modified glass substrates and active mixing enhanced the biosensing performance compared to the use of bare glass and simple diffusion. Finally, we performed functional secretory analysis of interleukin-8 and tumor necrosis factor alpha from human neutrophils and monocytes, stimulated with various doses of lipopolysaccharide and phorbol 12-myristate 13-acetate-ionomycin, respectively. We foresee the employment of our microfluidic platform in the diagnosis of different pathologies where alterations in cytokine secretion patterns can be used as biomarkers.
Keywords: MITOMI; cytokine secretion; fluorescence biosensors; immune cells; microfluidic immunoassays; single cells.