An ultrasensitive and conformation sensitive fluorescent probe for sensing human albumin in complex biological samples

Abstract

In this study, an ultrasensitive and conformation sensitive fluorescent probe HMM (2-((6-hydroxynaphthalen-2-yl) methylene) malononitrile) was developed for highly selective detection of human albumin (HA) in complex biological samples. Under physiological conditions, HMM could selectively bind on HA and form a complex which exhibited strong fluorescence signal around 562nm. Both Job’s Plot analysis and displacement assays using a series of site markers of HA demonstrated that HMM could simultaneously bind on two major ligand-binding sites (site I and site II) of HA, which partially explained why HMM could trigger more than 300-fold enhancement in fluorescence signal. The newly developed probe exhibited ultrahigh sensitivity (LOD=50ngmL−1) and excellent selectivity toward HA over other human serum proteins. HMM has been successfully used to detect HA in diluted human plasma and urine, and the results demonstrate that such detection is highly reliable. Unlike bromocresol green (a commonly used dye-binding probe of HA), HMM is a conformation sensitive fluorescent probe for HA and it could distinguish between native and denatured HA. All these features demonstrated that HMM could serve as a practical and powerful tool for HA associated clinical routine tests including quantification of serum and urinary albumin.