A poly(dimethylsiloxane)(PDMS)/SU-8/quartz hybrid chip was developed and applied in the isoelectric focusing (IEF) of proteins with ultraviolet (UV) absorbance-based whole-channel imaging detection (UV-WCID). Each hybrid chip was made of three layers: a PDMS flat top substrate, a bottom quartz substrate and a middle layer of SU-8 photoresist. The SU-8 serves two purposes: it contains the microchannel used for IEF separation, and acts as an optical slit that absorbs UV light below 300 nm improving detection sensitivity in WCID. The novel hybrid design demonstrates a two to three times improvement in sensitivity over a comparable PDMS/PDMS design. In addition, the hybrid chip exhibits increased heat dissipation due to the superior thermal conductivity of the bottom quartz substrate allowing for larger electric fields to be used in separations. The hybrid design with IEF-UV-WCID was successful in resolving a complicated sample, hemoglobin control, with high fidelity.