An Integrated Approach to Protein Discovery and Detection From Complex Biofluids

Ovarian cancer, a leading cause of cancer-related deaths among women, has been notoriously difficult to screen for and diagnose early, as early detection significantly improves survival. Researchers and clinicians seek routinely usable and noninvasive screening methods; however, available methods (i.e., biomarker screening) lack desirable sensitivity/specificity. The most fatal form, high-grade serous ovarian cancer, often originate in the fallopian tube; therefore, sampling from the vaginal environment provides more proximal sources for tumor detection. To address these shortcomings and leverage proximal sampling, we developed an untargeted mass spectrometry microprotein profiling method and identified cystatin A, which was validated in an animal model. To overcome the limits of detection inherent to mass spectrometry, we demonstrated that cystatin A is present at 100 pM concentrations using a label-free microtoroid resonator and translated our workflow to patient-derived clinical samples, highlighting the potential utility of early stage detection where biomarker levels would be low. [Display omitted] •A workflow was to identify high-grade serous ovarian cancer biomarkers•Mass spectrometry proteomics identified cystatin A•A surface conjugated antibody for cystatin A facilitated pM microtoroid detection High-grade serous ovarian cancer can originate in the fallopian tube epithelium. Tumors colonize the ovary and then metastasize throughout the peritoneum; however, no routine screening exists for routine health exams. We leveraged vaginal lavages from a murine model as a complex biological fluid for protein biomarker discovery. We discovered and validated cystatin A as a putative biomarker. Detection was improved using a cystatin A antibody conjugated to a microtoroid resonator’s surface, facilitating detection from vaginal lavages and patient-derived tampons.