Super-resolution Reflection Microscopy via Absorbance Modulation

Super-resolution Reflection Microscopy via Absorbance Modulation

In recent years, fluorescence microscopy has been revolutionized. Reversible switching of fluorophores has enabled circumventing the limits imposed by diffraction. Thus, resolution down to the molecular scale became possible. However, to the best of our knowledge, the application of the principles u...

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Bibliographic Details
Published in:ACS Nanoscience Au Vol. 3; no. 5; pp. 375 - 380
Main Authors: Jain, Parul, Geisler, Claudia, Leitz, Dennis, Udachin, Viktor, Nagorny, Sven, Weingartz, Thea, Adams, Jörg, Schmidt, Andreas, Rembe, Christian, Egner, Alexander
Format: Journal Article
Language:English
Published: American Chemical Society 18-10-2023
Subjects:
Letter
absorbance modulation imaging (AMI)
reflection microscopy
Nanoscopy
absorbance modulation layer (AML)
super-resolution
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Summary:In recent years, fluorescence microscopy has been revolutionized. Reversible switching of fluorophores has enabled circumventing the limits imposed by diffraction. Thus, resolution down to the molecular scale became possible. However, to the best of our knowledge, the application of the principles underlying super-resolution fluorescence microscopy to reflection microscopy has not been experimentally demonstrated. Here, we present the first evidence that this is indeed possible. A layer of photochromic molecules referred to as the absorbance modulation layer (AML) is applied to a sample under investigation. The AML-coated sample is then sequentially illuminated with a one-dimensional (1D) focal intensity distribution (similar to the transverse laser mode TEM01) at wavelength λ1 = 325 nm to create a subwavelength aperture within the AML, followed by illumination with a Gaussian focal spot at λ2 = 633 nm for high-resolution imaging. Using this method, called absorbance modulation imaging (AMI) in reflection, we demonstrate a 2.4-fold resolution enhancement over the diffraction limit for a numerical aperture (NA) of 0.65 and wavelength (λ) of 633 nm.
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ISSN:2694-2496
2694-2496
DOI:10.1021/acsnanoscienceau.3c00013