Protein biomarker detection via differential dynamic microscopy

We demonstrate a pioneering application of differential dynamic microscopy (DDM) for disease diagnosis by monitoring the dynamic signal of probe beads. When the target antibody binds to our sensitized probe beads, it allows these originally non-fluorescent beads to be indirectly decorated with fluorescently labeled secondary antibodies. Using standard epi-fluorescence microscopy without the need for purification, DDM reliably detects the resulting increase in fluorescence signal from the probe beads. It effectively discerns this signal from the background, which includes residual free fluorescent secondary antibodies. As a proof-of-concept, polystyrene beads (500 nm in diameter) were conjugated with the receptor binding domain of SARS-CoV-2’s spike protein to quantify immunoglobulin G1-specific antibodies against it in three different media of increasing complexity (phosphate buffer, model saliva and fetal bovine serum). Our results show that as the medium becomes more complex, more dilution and/or longer incubation time may be needed to overcome the competition effect between the target and interfering proteins. By optimizing imaging parameters and sample preparation we could surpass clinically relevant limits of detection in both saliva and serum samples. In contrast to conventional diagnostic techniques including ELISA and PCR, DDM brings many advantages including simplicity of the preparation protocol, high speed, and versatility without requiring very expensive or complicated equipment. As such, DDM-based disease diagnostics has the potential to be utilized in many laboratories, especially in remote regions with limited resources and point-of-use applications may also be feasible by miniaturizing the imaging setup.