Iodophenol blue-enhanced luminol chemiluminescence and its application to hydrogen peroxide and glucose detection.

In this study, we found that iodophenol blue can enhance the weak chemiluminescence (CL) of luminol-H2O2 system. With the aid of CL spectral, electron spin resonance (ESR) spectral measurements and studies on the effects of various free radical scavengers on the iodophenol blue-enhanced luminol-H2O2 system, we speculated that iodophenol blue may react with H2O2 and oxygen to produce oxidizing radical species such as OH(•) and O2(•-) resulting the formation of (1)O2. The generated (1)O2 may react with luminol anion generating an unstable endoperoxide and subsequent 3-aminophthalate* (3-APA*). When the excited-state 3-APA returned to the ground-state, an enhanced CL was observed. Based on the H2O2 concentration dependence of the catalytic activity of iodophenol blue, a cheap, simple, sensitive CL assay for the determination of H2O2 was established. Under the optimum experimental conditions, a linear relationship between the relative CL intensity and H2O2 concentration in the range of 0.025-10 µM was obtained. As low as 14 nM H2O2 can be sensitively detected by using the proposed method. The relative standard deviation for 5, 1 and 0.25 µM H2O2 was 2.58%, 5.16% and 4.66%, respectively. By combining the glucose oxidase (GOx)-catalyzed oxidation reaction, CL detection of glucose was realized. The linear range of glucose detection was 0.1-30 µM with a detection limit of 0.06 µM. The proposed method has been applied to the detection of glucose in diluted serum.

Turn-on chemiluminescent sensing platform for label-free protease detection using streptavidin-modified magnetic beads.

We report a label-free streptavidin-modified magnetic beads (SA-MBs)-based sensing platform for turn-on chemiluminescent (CL) detection of protease using trypsin as model analyte. In the assay, a biotinylated peptide containing an arginine and a terminal cysteine was used as the substrate of trypsin. Upon adding the peptide into a basic luminol-NaIO4 solution, the terminal cysteine induced a strong CL signal. Surprisingly a much lower CL was emitted when the peptide was immobilized on the surface of SA-MBs. Based on this phenomenon, we designed a turn-on CL sensing system for protease using trypsin as model and its inhibitors screening. In the absence of trypsin, the peptide was coupled to the SA-MBs surface, resulting in a low CL background. Upon the addition of trypsin, the peptide can be catalytically hydrolyzed at the C-terminus of arginine, resulting in the formation of free cysteine-containing residues and subsequent CL recovery with the addition of luminol and NaIO4. The simple method does not require washing or separating procedures. Trypsin at a concentration as low as 10 pM can be assayed using this new CL sensing system. Additionally, the proposed method can be employed for screening the inhibitors of trypsin. This new sensing strategy could be easily extended to assay other proteases by simply changing the peptide substrate.