Continuously Controlled Spectral-resolution (CoCoS) at High Resolution Confocal Imaging and Fast Scanning

Recently, super-resolution methods have been developed to improve the resolution and among them, techniques that combine super-resolution and wide-field imaging with structured illumination. These techniques are still diffraction-limited, but double the resolution of a microscope. One of these techniques is image scanning microscopy (ISM). It achieves high resolution by scanning a sample with a diffraction-limited illumination focus and records at each position a small image of the illuminated region. This technique is very time consuming, so combining ISM with confocal spinning disk (CSD) microscopy helps to increase image acquisition speed by ~100 times. This method is suited nicely for fast 3D super-resolution confocal imaging of fixed and live samples.

Our goal is to combine CSD-ISM microscopy with Continuously Controlled Spectral-resolution (CoCoS) microscopy- an imaging scheme that encodes color into spatial read-out in the image plane, with continuous control over the spectral resolution. Combining these methods will provide us the ability to record, detect and distinguish different entities located close to each other, usually at scales smatter than resolution limit of standard optical microscopy (~250 nm), at high resolution and fast scanning.

With this method, we will start with imaging a gel with multicolor 100nm beads in 3D. We will register the separate color images and align them according to the 3d bead map. Eventually, with this method we aim to observe fine tracking of drugs attaching to cell mitochondria.