Salivary Biomarkers and Microbiome Analysis

Salivary pH was digitally measured through a digital pH sensor (Fisher Scientific Accumet AE150). We initially calibrated the device using buffered solutions with pH 4.0, pH 7.0, and pH 10.0. After calibration, the sensor probe was dipped in saliva filled tube, where it remained for 30 s, thus yielding automatic pH reading. High sensitivity enzyme-linked immunosorbent sandwich assay kit (ELISA kit; Lifespan Biosciences LS-F27082) was used to determine the MUC7 levels in the saliva samples, following the protocol described by the manufacturer. Values of absorbance were read at the wavelength of A = 450 nm using the SpectraMax iD3 Multi-Mode Microplate Reader. Salivary MUC7 levels were estimated from standard curves, derived from the recombinant MUC7 standards. DNA copies of Streptococcus Mutans, Lactobacillus spp, and Candida Albcians were quantified via SYBR qPCR (Roche LightCycler480 Real Time PCR), using the primer sets and protocol described in previous studies [44 , 45 (link)].