A two-step competition assay for visual, sensitive and quantitative C-reactive protein detection in low-cost microfluidic particle accumulators

Abstract

Monitoring C-reactive protein (CRP) levels in point-of-care testing (POCT) scenarios is of great value in early diagnosis, prevention and treatment of inflammation in patient’s home, development country as well as other resource-limited settings. Here, we reported a sensitive, economic and simple biosensing approach for visual and sensitive CRP detection by trapping and accumulating microparticles in a photolithography-free, low-cost microfluidic chip. A competition assay among microparticles, magnetic nanoparticles and CRP antigen is utilized to quantify the CRP concentrations. When CRP antigen is present, fewer magnetic nanoparticles can bind to antibody-conjugated microparticles, leaving the free microparticles flowing and trapping in a microfluidic particle accumulation chip after magnetic separation, and forming a length-quantifiable visual bar, which is proportional to the concentrations of CRP in detection samples. Therefore, the target protein can be quantitatively detected by reading the length-quantifiable visual bar in a microfluidic channel by the naked eyes. Of note, a novel two-step competition assay was utilized to achieve the highly sensitive and specific CRP detection with a limit of detection (LOD) of ~ 32 pg/mL, and a wide detection range from 100 pg/mL to 10 µg/mL. The developed method also showed high selectivity, good accuracy and great stability to different temperature and incubation time. More importantly, the design of the sensor could be applicable for detecting any other biomarkers feasibly, thus providing a universal platform for visual, sensitive and economic clinical biomarker measurement by the naked eyes for POCT applications without any auxiliary equipment.