High-throughput microplate-based optical assay of salivary phosphates using image capturing detection

A high-throughput optical assay has been developed and validated for the direct determination of salivary phosphates. The assay is based on the adaptation of an advantageous version of the molybdenum blue reaction (using Sn(II)/hydrazine as the reductant) in a microplate platform combined with instrument-free detection using an overhead book scanner. Salivary phosphates could be determined in the absence of matrix interferences in the range of 10-100 µM following simple, 100-fold dilution of the biological samples. The precision of the assay was better than 2.1 and 6.4% within and between days, respectively, and the accuracy ranged between 88% and 120% (expressed as % recoveries at 20 and 40 µM phosphates). The mean phosphate concentration found in real human saliva was 4.4 ± 0.6 mM (n = 5), being in good agreement with values reported in the literature and corroborative analysis by an automated flow spectrophotometric method. The greenness of the procedure has also been evaluated using BAGI and GAPI tools.