Patterned illumination single molecule localization microscopy (piSMLM)

• Verfasst von: Chen, Shih-Ya [VerfasserIn]
• Verfasst von: Schaufler, Wladimir [VerfasserIn]
• Verfasst von: Cremer, Christoph [VerfasserIn]
• Titel: Patterned illumination single molecule localization microscopy (piSMLM)
• Titelzusatz: user defined blinking regions of interest
• Verf.angabe: Shih-Ya Chen, Felix Bestvater, Wladimir Schaufler, Rainer Heintzmann, and Christoph Cremer
• E-Jahr: 2018
• Jahr: [12 Nov 2018]
• Umfang: 12 S.
• Illustrationen: Illustrationen
• Fussnoten: Gesehen am 12.04.2019
• Titel Quelle: Enthalten in: Optics express
• Ort Quelle: Washington, DC : Soc., 1997
• Jahr Quelle: 2018
• Band/Heft Quelle: 26(2018), 23, Seite 30009-30020
• ISSN Quelle: 1094-4087
• DOI: doi:10.1364/OE.26.030009
• Datenträger: Online-Ressource
• Sprache: eng
• Sach-SW: Image quality
• Sach-SW: Liquid crystal modulators
• Sach-SW: Optical components
• Sach-SW: Optical design
• Sach-SW: Phase retrieval
• Sach-SW: Speckle noise
• K10plus-PPN: 1663203881

Abstract

Single molecule localization microscopy (SMLM) has been established as an important super-resolution technique for studying subcellular structures with a resolution down to a lateral scale of 10 nm. Usually samples are illuminated with a Gaussian shaped beam and consequently insufficient irradiance on the periphery of the illuminated region leads to artifacts in the reconstructed image which degrades image quality. We present a newly developed patterned illumination SMLM (piSMLM) to overcome the problem of uneven illumination by computer-generated holography. By utilizing a phase-only spatial light modulator (SLM) in combination with a modified Gerchberg-Saxton algorithm, a user-defined pattern with homogeneous and nearly speckle-free illumination is obtained. Our experimental results show that irradiance 1 to 5 kW/cm2 was achieved by using a laser with an output power of 200 mW in a region of 2000 µm2 to 500 µm2, respectively. Higher irradiance of up to 20 kW/cm2 can be reached by simply reducing the size of the region of interest (ROI). To demonstrate the application of the piSMLM, nuclear structures were imaged based on fluctuation binding-activated localization microscopy (fBALM). The super-resolution fBALM images revealed nuclear structures at a nanometer scale.