A Miniaturized Optical Bioassay Towards Early‐Stage Diagnosis of Cancer Using Immunosensing of Biomarkers (CA 15.3, PSA and CEA) by Toluidine Blue‐Decorated Poly(Methyl Methacrylate): Proof of Concept

The current study evaluated the application of a novel and advanced photochemical biosensing platform for the detection of multiple biomarkers of prostate-specific antigen (PSA), carcinoembryonic antigen (CEA) and cancer antigen 15-3 (CA 15.3) in human plasma samples using optical biomedical analysis. For the first time, toluidine blue (TB)-decorated polymethyl methacrylate (PMMA) micro-arrays integrated with smartphone-based digital image analysis towards cancer biomarkers detection. Immunosensing was achieved through antigen-antibody binding, which induce TB aggregation via immunocomplex formation, resulting in quantifiable colorimetric shifts mediated by surface plasmon resonance (SPR) modulation. The assay demonstrated linear ranges of 1-60, 1-75 and 1-125 µg/mL, with a low limit of quantification of 10, 1 and 1 µg/mL, for PSA, CEA and CA 15.3, respectively. A smartphone-enabled ambient light sensor was employed for real-time digital colorimetry, enabling user-friendly quantification of prostate, colon and breast cancer biomarker without specialized instrumentation. The developed system addresses critical gaps in point-of-care diagnostics by combining scalable PMMA microarray fabrication, targeted TB-antibody conjugation chemistry and machine vision-based signal processing into a unified workflow. This low-cost, multiplexed sensing approach holds significant promise for early cancer screening in resource-limited settings, with potential applications in clinical diagnostics, remote health monitoring and personalized medicine.