The cytokine production by leukocytes correlates with body's ability to mount an immune response and therefore has high diagnostic value. In the present study we employed microfabricated surfaces to capture T-cells from minimally processed human blood, arrange these cells into a single cell array, and then detect interferon (IFN)-γ released from individual cells. The fabrication of cell capture surfaces started with coating a silane-modified glass slide with a uniform layer of poly(ethylene glycol) (PEG) hydrogel. The hydrogel-coated slide was lyophilized and then incubated with a mixture of monoclonal anti-IFN-γ and anti-CD4 antibodies (Abs). To define sites for single cell attachment, PEG hydrogel microwells (20 μm diameter) were photolithographically patterned on top of the Ab-containing hydrogel layer. This micropatterning process resulted in fabrication of PEG hydrogel microwells with Ab-decorated bottom and nonfouling walls. To minimize the blood volume requirement and to precisely define shear stress conditions, the engineered surface was enclosed inside a PDMS-based microfluidic device. Introduction of red blood cell (RBC) depleted whole human blood followed by controlled washing led to the isolation of individual CD4 T-cells within PEG microwells. Mitogenic activation and immunofluorescent staining performed inside the microfluidic chamber revealed IFN-γ cytokine signal colocalized with specific T-cells. The device and process presented here will be expanded in the future to enable multiparametric functional analysis of immune cells organized into high density single cell arrays.
|Original language||English (US)|
|Number of pages||7|
|State||Published - Oct 1 2009|
ASJC Scopus subject areas
- Analytical Chemistry