Development of immunomagnetic droplet-based digital immuno-PCR for the quantification of prostate specific antigen

Abstract

Immuno-PCR (IPCR) is an immunoassay that employs DNA as the signal generator and utilizes both the versatility of antigen–antibody interactions and the exponential amplification power of PCR. Droplet-based digital PCR (ddPCR) is the third and latest generation of PCR technology, and the integration of ddPCR into IPCR will hopefully enable the absolute quantification and highly sensitive detection of proteins. However, the lack of practical washing methods in the microfluidic droplet system still remains a huge obstacle for such integration. Here, we report a novel droplet-based digital immuno-PCR (ddIPCR) method with magnetic beads as the solid phase. Supported by their separate functions with and without a magnetic field, the employment of magnetic beads successfully builds a bridge between IPCR and droplet microfluidics. Using prostate specific antigen (PSA) as a model antigen, the established immunomagnetic ddIPCR method could detect PSA down to 0.48 ng mL⁻¹ with a linear range from 0.5 to 30 ng mL⁻¹. Also, the detection of human serum samples by immunomagnetic ddIPCR correlated well with the clinical results. The immunomagnetic ddIPCR method provides a novel and highly sensitive detection route for the quantification of antigens, showing great potential in protein research and clinical diagnosis.