UC San Diego

Background: Pancreatic ductal adenocarcinoma (PDAC) is among the most lethal malignancies worldwide, with a five-year survival rate of approximately 12–13% due to the absence of reliable early detection tools. Elevated circulating protease activity, including cathepsin S and chymotrypsin, has been demonstrated in PDAC patient blood using the Rapid Assay for Protease Detection (RAPD), a charge-changing peptide (CCP)-based gel electrophoresis platform developed at UC San Diego. Dried blood spot (DBS) cards offer a cold-chain-independent sample blood sample collection from a finger prick that could enable home-based PDAC screening; however, no validated method exists for protease activity detection from DBS matrices.Result: This thesis tests the feasibility and storage stability of CCP-based protease activity detection from PDAC patient whole blood spotted onto blood spot cards (BSCs; Whatman 903 protein saver cards). Whole blood spotted onto BSCs produced significantly higher normalized fluorescent intensity than matched plasma for both cathepsin S (2.7-fold; p=0.030) and chymotrypsin (2.3-fold; p=0.003) substrates, consistent with the hypothesis that significant protease activity is present in the cellular fraction of whole blood. BSC storage temperatures between 4°C, –20°C, and –80°C revealed no statistically significant differences in protease activity for either substrate (cathepsin S: p=0.503; chymotrypsin: p=0.182 by one-way ANOVA), suggesting that refrigerator-temperature storage may be sufficient for clinical deployment.Conclusion: These findings provide a foundational pre-clinical validation for a BSC-based PDAC diagnostic system in which fingerstick blood samples can be mailed as cards into a centralized RAPD analysis, without phlebotomy or cold-chain logistics.