A supercharged fluorescent protein based FRET sensing platform for detection of heparin contamination

Abstract

Development of biosensors for the detection of oversulfated chondroitin sulfate (OSCS), the key contaminant in heparin (Hep), is a greatly demanding but challenging task. In this work, a FRET system for the detection of OSCS was successfully constructed by using supercharged green fluorescent protein (ScGFP) as the energy donor and dye labelled Hep (Hep-RF1) as the energy acceptor. With heparinase treatment, Hep-RF1 was hydrolyzed into small fragments, resulting in quenching of the FRET signal. On the other hand, since OSCS is an inhibitor for heparinase, the presence of OSCS would enable the effective FRET from ScGFP to Hep-RF1 even after heparinase treatment. With this ScGFP based FRET sensing platform, as low as 0.001% (w/w) OSCS in Hep has been successfully detected.