Novel Electrochemical Methodology for Activity Estimation of Alkaline Phosphatase Based on Solubility Difference

Abstract

We propose a novel electrochemical detection system for alkaline phosphatase (ALP) activity using the difference in water and oil solubilities between the substrate, ferrocene ethyl phosphate ester (FcEtOPO(3)(2-)), and the enzymatic product, ferroceneethanol (FcEtOH). In this system, water droplets containing ALP and FcEtOPO(3)(2-) were placed on a Pt disk microelectrode and surrounded by a mineral oil. By the ALP-catalyzed reaction, FcEtOPO(3)(2-) was converted to FcEtOH, which was then transferred to the mineral oil from the water droplets with FcEtOPO(3)(2-) remaining in the water droplets. After partitioning FcEtOH from the water droplets, FcEtOPO(3)(2-) was detected at the Pt disk microelectrode to estimate the ALP activity. Using this novel system, the ALP activity of embryoid bodies was successfully detected. We believe that the present system will be widely applicable to ALP-based bioassays.

Bifunctional magnetic nanobeads for sensitive detection of avian influenza A (H7N9) virus based on immunomagnetic separation and enzyme-induced metallization

“…The amount of ALP was estimated by the p-NPP method (Ino et al, 2012) since ALP can catalyze p-NPP to produce p-nitrophenol, the details could be found in Supporting information. The reaction between the bi-MBs and p-NPP produced a yellow solution which indicated the successful modification of ALP on biMBs surface.…”

Nitrogen-doped Carbon Dots Mediated Fluorescent on-off Assay for Rapid and Highly Sensitive Pyrophosphate and Alkaline Phosphatase Detection

“…Up to date, many techniques have been developed for the detection of PPi and ALP including colorimetric 10, 11 , fluorometric 12, 13 , surface-enhanced Raman scattering 14 and electrochemical methods 15, 16 . Among these assays, fluorometric methods have attracted considerable interest for their rapid response, easy operation and high sensitivity 17–19 .…”

Microchemistry- and MEMS-based Integrated Electrochemical Devices for Bioassay Applications

“…The development of several types of electrochemical devices for bioassay applications, including analyses of yeast and mammalian cells, has been reported. [29][30][31][32][33][34][35][36][37][38][39][40][41][42][43][44] IDA electrodes were incorporated to amplify signals from cells and provide highly sensitive assays. Yeast cellbased IDA electrodes were utilized for the detection of hormone active chemicals.…”