Abstract
.Significance: It is commonly assumed that using the objective lens to create a tightly focused light spot for illumination provides a twofold resolution improvement over the Rayleigh resolution limit and that resolution improvement is independent of object properties. Nevertheless, such an assumption has not been carefully examined. We examine this assumption by analyzing the performance of two super-resolution methods, known as image scanning microscopy (ISM) and illumination-enhanced sparsity (IES).Aim: We aim to identify the fundamental differences between the two methods, and to provide examples that help researchers determine which method to utilize for different imaging conditions.Approach: We input the same image datasets into the two methods and analyze their restorations. In numerical simulations, we design objects of distinct brightness and sparsity levels for imaging. We use biological imaging experiments to verify the simulation results.Results: The resolution of IES often exceeds twice the Rayleigh resolution limit when imaging sparse objects. A decrease in object sparsity negatively affects the resolution improvement in both methods.Conclusions: The IES method is superior for imaging sparse objects with its main features being bright and small against a dark, large background. For objects that are largely bright with small dark features, the ISM method is favorable.
Highlights
Using a tightly focused light spot for illumination has been one of the most significant advancements in the history of optical microscopy
Using the synthetic image data generated by the above simulations, we compare the restorations of the two methods as we vary the image signal-to-noise ratio (SNR) (Fig. 2) and object sparsity (Fig. 3)
For the illumination-enhanced sparsity (IES) method, by contrast, Donoho et al argued that, as long as the image SNR is sufficient, objects of high sparsity can always be resolved by non-negative least-squares (NNLS) deconvolution, the required SNR may be prohibitively high in practice.[17]
Summary
Using a tightly focused light spot for illumination has been one of the most significant advancements in the history of optical microscopy. Yu et al.: Analyzing the super-resolution characteristics of focused-spot illumination. Scanning microscopy (ISM),[10] due to its extraordinary photon efficiency as well as its confocal super-resolution effect.[11]. The original ISM method uses the objective lens to create a tightly focused illumination spot, steps this spot across the object, and acquires one descanned image at each step.[10] For images acquired without a descan arrangement, each acquired image is cropped with respect to its illumination spot position.[12] For these descanned or cropped images, each pixel is considered equivalent to a pinhole detection in a confocal microscope and is reassigned to create a particular confocal image [Fig. 1(a)]. Many confocal images are generated from a complete lateral scan
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