Paper Analytical Device for the Colourimetric Detection of Alkaline Phosphatase in Serum and Saliva

A paper analytical device (PAD) was developed for the colourimetric estimation of alkaline phosphatase (ALP). ALP catalyzes the hydrolysis of disodium phenyl phosphate (DSPP) to phenol, which then undergoes oxidative coupling with 4-aminoantipyrine (4-AAP) producing a reddish brown-coloured quinone imine. The colourimetric reaction tested in solution has been translated onto a PAD for the point of care (PoC) testing of ALP. A uniform colour was obtained in 25 min with the introduction of ALP onto the reagent drop cast paper device. The images obtained by scanning the PAD were processed using ImageJ software. The colour intensity obtained by image processing was proportional to the concentration of ALP present in the sample. ALP in serum and saliva samples were tested using the PAD in the range of 0 to 528 U/L and 0 to 187 U/L, respectively. The interference studies revealed that the PAD was selective to ALP in the presence of other biomolecules. The PAD exhibited a limit of detection (LOD) of 3.3 U/L and a sensitivity of 48.36 a.u/Log(U/L) for serum ALP and LOD of 1.69 U/L and a sensitivity of 28.5 a.u/Log (U/L) for salivary ALP. The PAD showed good agreement with the clinical methods for real samples.