We report methods for stabilizing cellulose-based immunoassays and using this platform to analyze human saliva. Stabilization treatments of immunoassays for matrix metalloproteinases (MMP)-8 and -9, biomarkers of periodontal disease, were conducted and compared, revealing that anti-MMP-8 and -9 capture antibodies could be stabilized with the addition of a 5% trehalose solution to the test zones, followed by drying in a vacuum oven. After stabilization, the paper devices retained equivalent binding activity to that of freshly prepared tests for 14 days a time frame that enables US-based clinical testing of this diagnostic assay. A saliva pretreatment method was developed to remove viscous elements without reducing the concentration or binding activity of dissolved proteins. Immunoassays were stored in ziplock bags containing desiccant, and used to detect nanomolar concentrations of MMP-9 in human saliva across the relevant clinical concentration range. These methods and findings facilitate rapid, affordable validation studies of this and other biomarkers that are found in saliva using vertical flow immunoassays.
Why is this work important?
Low-cost detection of biomarkers associated with the majority of diseases is not feasible.
What has been done before?
Paper-based assays have been used to detect biomarkers in human serum or blood. The paper-based detection of biomarkers in saliva has not been extensively studied.
What are our contributions?
We propose methods for stabilizing paper-based immunoassays for robust and low-cost detection of salivary biomarkers. This method is low-cost and can facilitate rapid detection of oral biomarkers. We specifically describe the detection of biomarkers associated with oral diseases, MMP-8 and -9.
What are the next steps?
We are expanding this approach to assay biomarkers in serum and blood.