A ready-to-use, versatile, multiplex-able three-dimensional scaffold-based immunoassay chip for high throughput hepatotoxicity evaluation.

Hydrogel as three-dimensional (3D) substrate has been employed in miniaturized high throughput protein detection platforms to increase the number of effective antibodies and signal augmentation. However, the high water content of the hydrogel can dilute samples and create barrier to mass transfer, limiting hydrogel height to several microns in most platforms. Moreover, these platforms cannot achieve widespread use in common laboratories as they usually rely heavily on expensive robotic liquid handlers and custom-built components. Here we developed a ready-to-use, easy to store and handle, versatile and multiplex-able 3D scaffold-based immunoassay chip (3D immunoChip) possible for high throughput protein quantification using bench-top equipment in common laboratories. Sample dilution, mass transfer, signal scattering and storage problems can be avoided by using dry scaffolds that regain transparency upon rehydration. When combined with hydrophilic-hydrophobic patterned reagent loading slides, manual high throughput handling of samples can be achieved. As these micro-scaffolds are patterned without barriers in between, simultaneous and effortless washing of all the reaction zones is possible in a Petri dish. Such features aid the 3D immunoChip in saving up to 100 times reagent and about 6 times labour. The 3D immunoChip is able to detect albumin (ALB), as a model analyte, from 5 ng mL(-1) to 1000 ng mL(-1), making it comparable to the commercialized ELISA kit based on a 96-well plate (0.22-400 ng mL(-1)). This thus enables the 3D immunoChip to directly detect ALB secreted by HepaRG cells cultured in a 3D cell culture array chip for high throughput drug hepatotoxicity evaluation, which could potentially accelerate drug screening.