Steric Hindrance Assay for Secreted Factors in Stem Cell Culture

The ex vivo expansion of hematopoietic stem cells is significantly inhibited by secreted proteins that induce negative feedback loops. The ability to effectively monitor these factors is critical for their real-time regulation and control and, by extension, enhancing stem cell expansion. Here, we describe a novel monitoring strategy for the detection of soluble signaling factors in stem cell cultures using a DNA-based sensing mechanism on a chip-based nanostructured microelectrode platform. We combine DNA hybridization engineering with antibody-capturing chemistry in an amplified steric hindrance hybridization assay. This method enables the quantification of important secreted proteins, showcased by the detection of 10 pg·mL^-1 level concentrations of three proteins in stem cell culture samples. This approach is the first universal nonsandwich technique that permits pg·mL^-1 level quantification of small proteins in stem cell culture media without signal loss.