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

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 .