Smartphone‐Based Free‐to‐Total Prostate Specific Antigen Ratio Detection System Using a Colorimetric Reaction Integrated with Proximity‐Induced Bio‐Barcode and CRISPR/Cas12a Assay

Abstract The free‐to‐total prostate‐specific antigen (f/t‐PSA) ratio is of great significance in the accurate diagnosis of prostate cancer. Herein, a smartphone‐based detection system is reported using a colorimetric reaction integrated with proximity‐induced bio‐barcode and the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas12a assay for f/t‐PSA ratio detection. DNA/antibody recognition probes are designed to bind f‐PSA or t‐PSA and induce the release of the DNA bio‐barcode. The CRISPR/Cas12a system is activated by the DNA bio‐barcode to release Ag+ from the C‐Ag+‐C structure of the hairpin DNA. The released Ag+ is used to affect the tetramethylbenzidine (TMB)‐H2O2‐based colorimetric reaction catalyzed by Pt nanoparticles (NPs), as the peroxidase‐like activity of the Pt NPs can be efficiently inhibited by Ag+. A smartphone with a self‐developed app is used as an image reader and analyzer to analyze the colorimetric reaction and provide the results. A limit of detection of 0.06 and 0.04 ng mL −1 is achieved for t‐PSA and f‐PSA, respectively. The smartphone‐based method showed a linear response between 0.1 and 100 ng mL −1 of t‐PSA or f‐PSA. In tests with clinical samples, the smartphone‐based method successfully diagnosed prostate cancer patients from benign prostatic hyperplasia patients and healthy cases with high sensitivity and specificity.