Currently, it remains challenging to count protein-biomarker molecules present in a small droplet of biological samples. Herein, we propose a gold nanoparticle (GNP) probe-assisted sandwich-counting strategy that relies on a GNP probe, an antibody-functionalized chip to "count" antigen molecules using a scanning electron microscope. Both standard carcinoembryonic antigen (CEA) and two real CEA-related tumor samples (tumor tissues and serum) were assayed to demonstrate the proof-of-concept of the counting strategy. Results show that our method is excellently correlative with enzyme-linked immuno-sorbent assay (ELISA) that is widely used in clinics for antigen or antibody detection and the limit of detection of our enumeration strategy reaches down to 0.045 ng/mL, which is ∼40 times more sensitive than the conventional ELISA. Therefore, our GNP probe-assisted sandwich-counting strategy has the potential to be used for quantification of protein biomarkers at ultralow concentrations in early tumor specimens and detection of target proteins in much diluted concentrations.
Keywords: carcinoembryonic antigen; counting chip; gold nanoparticle probes; protein biomarker; scanning electron microscope (SEM).