A SIMPLE AND RAPID IMMUNOASSAY SYSTEM USING GREEN FLUORESCENT PROTEINTAG

A fusion protein between green fluorescent protein (GFP) and neuron-sp ecific enolase (NSE) was expressed in Escherichia coli. The GFP-NSE fu sion protein migrated at 62 kDa in SDS-PAGE and retained the fluoresce nce under non-heating conditions. However, heat-denatured GFP-NSE was non-fluorescent and migrated at 74 kDa corresponding to the theoretica l value. This suggests that the special structure of GFP, which is not denatured by SDS, influences its mobility in SDS-PAGE under non-heati ng conditions. The fluorescence intensity of GFP-NSE was measurable ov er a wide range by spectrophotometry or densitometry. The competitive immunoassay for NSE was performed using GFP-NSE as labeled antigen. Un der our assay conditions, the working range of this system was about 2 - 60 ng. This simple and rapid fluorescence immunoassay (FlA) using G FP-tagged antigen may be applicable to many protein markers.