Illumina Cambridge Limited

Illumina Corporation

Gary Mark Skinner

Stanley S Hong

Geraint Wyn Evans

Jonathan Mark Boutell

John A Moon

M Shane Bowen

Hongji Ren

Alex Nemiroski

Kenny Chen

Peter Clarke Newman

Simon Prince

Merek C Siu

Jason Richard Betley

Danilo Condello

Shaoping Lu

Aaron Liu

Gareth Jenkins

An imaging system includes: a sample container including a flowcell that includes a plurality of attachment elements at a plurality of sample locations, wherein a single DNA template molecule is attached to each of the attachment elements, and further wherein an average distance between adjacent attachment elements is less than Abbe's limit; and an imager positioned to image photo-switching occurring at the plurality of attachment elements by capturing on and off events in a plurality of color channels at the same time as the on and off events are occurring for the attached molecules when a stochastic photo-switching chemistry is applied to all of the attached molecules at the same time causing the attached molecules to fluoresce in the on and off events in up to four different colors.

Generalized stochastic super-resolution sequencing

In an example of the method, a functionalized coating layer is applied in depressions of a patterned flow cell substrate. The depressions are separated by interstitial regions. A primer is grafted to the functionalized coating layer to form a grafted functionalized coating layer in the depressions. A hydrogel is applied on at least the grafted functionalized coating layer.

Flow cells with hydrogel coating

Techniques are described for reducing the number of angles needed in structured illumination imaging of biological samples through the use of patterned flowcells, where nanowells of the patterned flowcells are arranged in, e.g., a square array, or an asymmetrical array. Accordingly, the number of images needed to resolve details of the biological samples is reduced. Techniques are also described for combining structured illumination imaging with line scanning using the patterned flowcells.

Reduced dimensionality structured illumination microscopy with patterned arrays of nanowells

The disclosure provides for structured illumination microscopy (SIM) imaging systems. In one set of implementations, a SIM imaging system may be implemented as a multi-arm SIM imaging system, whereby each arm of the system includes a light emitter and a beam splitter (e.g., a transmissive diffraction grating) having a specific, fixed orientation with respect to the system's optical axis. In a second set of implementations, a SIM imaging system may be implemented as a multiple beam splitter slide SIM imaging system, where one linear motion stage is mounted with multiple beam splitters having a corresponding, fixed orientation with respect to the system's optical axis. In a third set of implementations, a SIM imaging system may be implemented as a pattern angle spatial selection SIM imaging system, whereby a fixed two-dimensional diffraction grating is used in combination with a spatial filter wheel to project one-dimensional fringe patterns on a sample.

Dual optical grating slide structured illumination imaging

A method of sequencing a plurality of polynucleotides includes: attaching a single DNA template molecule to each of a plurality of attachment elements on a sample container, wherein the average distance between adjacent elements is less than Abbe's limit; applying a stochastic photo-switching chemistry to all of the molecules at the same time to cause the attached molecules to fluoresce in on and off events in up to four different colors by stochastic photo-switching; and imaging the on and off events in a color channel for each color in real-time as the on and off events are occurring for the attached molecules.

Example super-resolution microscopy systems are described herein that are configured for relatively high throughput. The disclosed microscopy systems can be to generate an array of sub-diffraction activated areas for imaging. The microscopy systems can be to utilize imaging techniques that employ time delay integration to build up super-resolution images over time. The disclosed microscopy systems can utilize long-lived fluorophores in conjunction with wide field and patterned illumination to generate super-resolution images of a sample with relatively high throughput.

Super-resolution microscopy

Presented herein are methods and compositions for enhancing utilization of surface primers during the surface amplification process. The methods are useful for surface amplification at improved densities. The methods and compositions provided herein enable creation of clusters which are brighter, but at the same densities as currently achieved using standard cluster amplification.

Enhanced utilization of surface primers in clusters

An apparatus and method for applying a chemistry to samples of interest are provided. The apparatus and method include a flexible tape mounted on an arrangement of guide rollers. Samples of interest (e.g., clusters of DNA templates) are bound to at least one surface of the flexible tape. The method and apparatus further comprise one or more read heads in relation to the flexible tape and a plurality of reservoirs along a path of the flexible tape. The reservoirs comprise liquids comprising chemical reagents for performing the chemistry on the samples of interest bound to the at least one surface of the flexible tape. The method and apparatus further comprise a drive system for driving at least one of the guide rollers to advance the flexible tape into and out of the reservoirs.

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Kedington, United Kingdom

Gary Mark Skinner