In this study we demonstrate that hydrogels loaded with a concentration gradient of an active molecule can be used as analytical tools for high throughput screening (HTS) and drug discovery applications. We used a microfluidic mixer to fabricate poly(ethylene glycol) (PEG) hydrogels that contained chemical concentration gradients. Each hydrogel was placed in culture medium in the vicinity of preosteoblast MC3T3-E1 cell layers. The controlled release of the toxin from the hydrogel in a gradient manner was used to study the effect of various chemical concentrations on cell viability. Given its simplicity, ease of storage and portability, this platform is a potentially useful approach to screen a range of drug concentrations simultaneously.