Wide-field interferometric microscopy is a common-path interferometry technique that allows for label-free and high-throughput detection of weakly scattering sub-diffraction-limited biological nanoparticles. Such nanoparticles appear as diffraction-limited-spots in the image and optically resolving them beyond their ‘digital’ detection still remains a challenge owing to the diffraction barrier as well as the typical signal levels that fall below the noise floor. In this study, we demonstrate the utility of computational optics in the interference enhanced nanoparticle imaging to improve its resolving power to obtain structural information on clinically relevant and often complexed-shaped biological nanoparticles such as viruses and exosomes. We consider a spatially incoherent structured illumination based image reconstruction strategy in wide-field interferometric microscopy to achieve high contrast nanoparticle imaging with super-resolution. Our reconstruction technique makes use of the optical transfer function of the system derived via an analytical model based on angular spectrum representation. We provide experimental demonstrations using an artificial sample to quantify the resolution enhancement as well as a biological sample for concept demonstration. We also benchmark the results against gold standard images obtained using an electron microscope. Our highly-sensitive super-resolution imaging system constitutes a noncomplex optical design, which can be realized with simple modifications to a conventional epi-illumination microscope, offering a cost-effective alternative to the laborious and expensive standard high-resolution microscopy techniques. It has a broad spectrum of applications ranging from clinical diagnostics to biotechnological research.
Current state-of-the-art technology for in-vitro diagnostics employ laboratory tests such as ELISA that consists of a multi-step test procedure and give results in analog format. Results of these tests are interpreted by the color change in a set of diluted samples in a multi-well plate. However, detection of the minute changes in the color poses challenges and can lead to false interpretations. Instead, a technique that allows individual counting of specific binding events would be useful to overcome such challenges. Digital imaging has been applied recently for diagnostics applications. SPR is one of the techniques allowing quantitative measurements. However, the limit of detection in this technique is on the order of nM. The current required detection limit, which is already achieved with the analog techniques, is around pM. Optical techniques that are simple to implement and can offer better sensitivities have great potential to be used in medical diagnostics. Interference Microscopy is one of the tools that have been investigated over years in optics field. More of the studies have been performed in confocal geometry and each individual nanoparticle was observed separately. Here, we achieve wide-field imaging of individual nanoparticles in a large field-of-view (~166 μm × 250 μm) on a micro-array based sensor chip in fraction of a second. We tested the sensitivity of our technique on dielectric nanoparticles because they exhibit optical properties similar to viruses and cells. We can detect non-resonant dielectric polystyrene nanoparticles of 100 nm. Moreover, we perform post-processing applications to further enhance visibility.