ABSTRACT
One major limitation of labeling proteins with synthetic fluorophores is the high fluorescence background, which necessitates extensive washing steps to remove unreacted fluorophores. In this paper, we describe a novel fluorogenic probe based on an environment-sensitive fluorophore for labeling with SNAP-tag proteins. The probe exhibits dramatic fluorescence turn-on of 280-fold upon being labeled to SNAP-tag. The major advantages of our fluorogenic probe are the dramatic fluorescence turn-on, ease of synthesis, high selectivity, and rapid labeling with SNAP-tag. No-wash labeling of both intracellular and cell surface proteins was successfully achieved in living cells, and the localization of these proteins was specifically visualized.
Subject(s)
Cytosol/metabolism , Fluorescent Dyes/chemistry , Molecular Imaging/methods , Molecular Probes , Proteins/metabolism , Flow Cytometry , HCT116 Cells , HumansABSTRACT
In this communication, we report a simple albumin probe based on a fluorescent molecular rotor for the detection of trace albumin levels in urine. In the presence of albumin, the probe exhibits remarkable 400-fold fluorescence enhancement with high selectivity and sensitivity. The probe was successfully applied in the quantitative detection of urinary albumin.
Subject(s)
Fluorescent Dyes/chemistry , Serum Albumin/analysis , Spectrometry, Fluorescence , Humans , Protein BindingABSTRACT
A general approach to mimic the sensing scheme of allosteric enzymes is developed. Through the covalent labeling of a sulfonamide inhibitor to the enzyme HCAII via SNAP-tag protein, the enzyme is rendered inactive. Catalytic activation is triggered only when a protease is present to cleave the recognized peptide sequence.
