Colorimetric Detection of Alkaline Phosphatase on Microfluidic Paper-based Analysis Devices

A low-cost, simple and sensitive colorimetric detection method for alkaline phosphatase (ALP) was developed on microfluidic paper-based analysis devices. In a typical colorimetric detection, 5-bromo-4-chloro-3-indolyl phosphate disodium salt (BCIP) and nitro blue tetrazolium (NBT) were firstly added onto the circle array zones of paper devices. Then ALP solutions were spotted to the array zones to perform the colorimetric reaction. The color reaction results were recorded by both Gel Documentation systems and a common digital camera, and analyzed with Quantity One software. All the reaction conditions were optimized. Under the optimal conditions, the colorimetric intensity showed a linear correlation to the concentration of ALP in the range of 1.5-20 U L-1 with a limit of detection (LOD) of 0.78 U L-1 (3s), which was about two orders of magnitude lower than that of the reported method on mu PADs. Besides, it was applied in the analysis of spiked real sample with a satisfactory result. In addition, the dual-color array-based paper strip was fabricated for the semiquantitative detection of ALP. The approximate activity of ALP could be simply distinguished by the color change with naked eyes.