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Super resolution microscopy techniques allows light microscopes to image details at extreme resolutions. However, as the resolution get better - the imaged area/volume gets smaller. For single-molecule localization microscopy this typically results in a FOV of around 50 um x 50 um. Chip NanoImaging (CNI) is introducing chip-based TIRF illumination, harnessing the powers of photonic integrated circuits. Using CNI technology, the illumination is decoupled from the microscope, which allows ultra-large FOV TIRF imaging. The resolution of the image depend on the imaging method used - from diffraction limited to super-resolution, all with 100 x larger field of view than current state-of-the-art.

The power of nanoscopy

Most super-resolution techniques today are based on fluorescence microscopy. Super-resolution microscopy is a relatively new technology that makes it possible to see small details that are impossible to see with traditional optical microscopy. Our contribution to nanoscopy is the introduction of a novel chip concept comprising a waveguide that allows for better illumination of a much larger area compared to existing technology.

Chip NanoImaging super-resolution microscopy demonstrated on a liver cell stained for F-actin.

Chip NanoImaging super-resolution microscopy demonstrated on a liver cell stained for F-actin.

Easily switch between EPI and TIRF illumination using ACP-ZERO

Epifluorescence (EPI) illuminates the entire sample, exciting fluorophores throughout the volume. While Total Internal Reflection Fluorescence (TIRF) selectively excites fluorophores close to the surface, resulting in high contrast and reduced background fluorescence.

Schematic illustrates the difference between EPI and TIRF illumination. EPI (right) illuminates the entire volume of the sample and excites all fluorescent molecules. The TIRF illumination (left) selectively illuminates fluorescent molecules within the exponentially decaying evanescent wave. A membrane stain demonstrates the difference between EPI (right) and TIRF (left) illumination.

Schematic illustrates the difference between EPI and TIRF illumination. EPI (right) illuminates the entire volume of the sample and excites all fluorescent molecules. The TIRF illumination (left) selectively illuminates fluorescent molecules within the exponentially decaying evanescent wave. A membrane stain demonstrates the difference between EPI (right) and TIRF (left) illumination.

PRINCIPLE OF OPERATION

ACP ZERO specialises in accessible and robust TIRF imaging. Our multimode waveguide technology decouples the illumination and imaging pathway. The imaging path follows conventional standards, while the illumination path occurs via waveguides patterned on the surface of a photonic chip. There are several waveguides in parallel on top of the chip, allowing for imaging within an active waveguide while preserving neighbouring fluorophores. The sample is placed or grown directly on top of the chip and laser light is automatically coupled into the waveguide. The evanescent field from the waveguide penetrates the sample and excites the fluorophores which are then detected by an upright microscope.

Photonic chip to illuminate the sample

Chip-based microscopy uses photonic integrated circuits as means to illuminate the sample. This means that the illumination can be tailored for the application, e.g., illuminating large areas using wide waveguides or illuminating pre-determined areas using waveguide branches like shown in the picture below with the star shaped waveguides.

Wafer with multiple chip sizes.

Picture showing chip with star shaped waveguides for research in SIM microscopy.

 

High-resolution  large content nanoimaging

Chip NanoImaging waveguide technology allows wide field of view illumination. The illuminated region is confined by the waveguide and can be made almost arbitrarily wide. The user is free to choose any magnification/NA objective lens for imaging. Crisp images with or without super-resolution.

 

Comparison of EPI (left) and TIRF (right) using different objectives. ChipNano TIRF allows the user to freely change the objective while maintaining a uniform TIRF illumination. Cells were labeled using F-actin AF647. All scalebars are 100 μm.

Comparison of EPI (left) and TIRF (right) using different objectives. ChipNano TIRF allows the user to freely change the objective while maintaining a uniform TIRF illumination. Cells were labeled using F-actin AF647. All scalebars are 100 μm.

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