Enzyme-triggered click chemistry combined with surface-enhanced Raman spectroscopy for the simple and sensitive detection of alkaline phosphatase activity from complex biological samples.

Alkaline phosphatase (ALP) is a widely used indicator in the diagnosis of various diseases. Thus, it is also urgent to develop simple and efficient methods, which can meet the accurate determination of ALP activity in physiological environments. In this study, enzyme-triggered click chemistry combined with the surface-enhanced Raman spectroscopy (SERS) technique was developed for the highly sensitive detection of the ALP activity in complex biological samples. ALP was able to catalyze the ascorbic acid-phosphate (AAP) to generate ascorbic acid (AA). Then, AA could reduce Cu(II) to produce Cu(I), which plays the role of a catalyst to promote the click reaction of azide terephthalic acid (ATA) and 4-acetylene biphenyl (4-AB), resulting in the SERS signal intensity of free 4-AB in the solution was clearly reduced along with the click reaction carried on, and showed a quantitative relationship with the concentration of ALP. The proposed method has the advantages of high sensitivity, selectivity and excellent repeatability. As a proof of concept, the new developed SERS-click strategy was applied to the specific determination of the ALP activity in clinical serum samples, cellular lysate samples and the ALP inhibitor assessment successfully, indicating that the proposed ALP-triggered click chemistry assay has a significant potential application in medical diagnosis.