Detection of Salivary Oral Cancer Biomarkers by Electrochemical Immunosensor Array

Objective: Interleukin-8 and -1b proteins in the cell-free phase of saliva have been identified as markers for oral cancer detection. Combined with electrochemical detection, these markers promise early identification and screening for disease through non-invasive procedures in a point-of-care setting. In this proof of principle study we present the development, optimization and application of two electrochemical immunoassays for the detection of IL-8 and IL-1b proteins directly from unprocessed human saliva.

Methods: Development started with buffer and reagent selection tests for a two-antibody sandwich assay with electrochemical detection on a sensor array. Antibody concentrations were then optimized to saliva protein levels based on linearity (R2) of standard curve, limit of detection (blank+2×StdDev), and coefficient of variation. After optimization, both electrochemical immunoassays were tested with protein standards and protein standard in saliva-spiked buffer. Finally, 11 oral cancer and 11 control clinical samples were selected to compare quantification to Luminex assay data.

Results: The protein electrochemical assays were able to discriminate between diseased and control groups with t-test p-value < 0.001. Data from the immunoassays showed strong correlation (R2=.92) with Luminex data.

Conclusion: Our data provided proof of concept data that the electrochemical protein detection platform can discriminate clinical saliva samples from oral cancer and healthy subjects. Our electrochemical immunoassays are inexpensive, fast and only needs 0.6 µl of saliva sample per reading. It puts us one important step closer to an innovative saliva based point-of-care detection of oral cancer.

Supported by PHS grant 5U01DE017790.