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Frontiers of Optoelectronics

Front. Optoelectron.    2015, Vol. 8 Issue (2) : 152-162     DOI: 10.1007/s12200-015-0479-x
REVIEW ARTICLE |
Resolution and contrast enhancements of optical microscope based on point spread function engineering
Yue FANG,Cuifang KUANG,Ye MA,Yifan WANG,Xu LIU()
State Key Laboratory of Modern Optical Instrumentation, Department of Optical Engineering, Zhejiang University, Hangzhou 310027, China
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Abstract

Point spread function (PSF) engineering-based methods to enhance resolution and contrast of optical microscopes have experienced great achievements in the last decades. These techniques include: stimulated emission depletion (STED), time-gated STED (g-STED), ground-state depletion microscopy (GSD), difference confocal microscopy, fluorescence emission difference microscopy (FED), switching laser mode (SLAM), virtual adaptable aperture system (VAAS), etc. Each affords unique strengths in resolution, contrast, speed and expenses. We explored how PSF engineering generally could be used to break the diffraction limitation, and concluded that the common target of PSF engineering-based methods is to get a sharper PSF. According to their common or distinctive principles to reshape the PSF, we divided all these methods into three categories, nonlinear PSF engineering, linear PSF engineering, and linear-based nonlinear PSF engineering and expounded these methods in classification. Nonlinear effect and linear subtraction is the core techniques described in this paper from the perspective of PSF reconstruction. By comparison, we emphasized each method’s strengths, weaknesses and biologic applications. In the end, we promote an expectation of prospective developing trend for PSF engineering.

Keywords super-resolution      optical imaging      point spread function (PSF) engineering      non-linear effects      linear subtraction     
Corresponding Authors: Xu LIU   
Issue Date: 24 June 2015
 Cite this article:   
Yue FANG,Cuifang KUANG,Ye MA, et al. Resolution and contrast enhancements of optical microscope based on point spread function engineering[J]. Front. Optoelectron., 2015, 8(2): 152-162.
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http://journal.hep.com.cn/foe/EN/10.1007/s12200-015-0479-x
http://journal.hep.com.cn/foe/EN/Y2015/V8/I2/152
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Yue FANG
Cuifang KUANG
Ye MA
Yifan WANG
Xu LIU
Fig.1  (a) Schematic image of a point-like object (left) and two close point-like objects (right); (b) image of point-like objects of (a) taken with a conventional (confocal) microscopy; (c) targeted image of point-like objects of (a); (d) conventional (confocal) light microscope’s PSF’s profile and targeted PSF’s profile
Fig.2  Profiles (1)-(4) show the spatial region in which the molecules are allowed to be state A (fluorescent state), if the region is illuminated with a doughnut-shaped patterns focused by a standing wave of light which drives molecules from A (fluorescent state) to B (non-fluorescent state) with peak intensities I 0 = 10 I sat , 50 I sat , 100 I sat and 500 I sat respectively whose corresponding cross section profiles are (i)-(iv) [ 30]
Fig.3  (a) Energy diagram of STED microscopes; (b) schematic representation of a STED microscope. A phase modulation mask is used to create a doughnut-shaped depletion beam overlapping the excitation laser beam
Fig.4  (a) Schematic representation of a difference confocal microscopy; (b) PSF profiles of conventional (large hole) microscopy system, confocal microscopy system and the subtractive result with α of 1/2
Fig.5  Principle of linear subtraction of a doughnut PSF from a solid PSF. (a) Solid PSF pattern; (b) doughnut-shaped PSF pattern by a 2π phase modulation; (c) subtractive of the doughnut-shaped PSF from the solid PSF with a proper factor; (d) profile curves of the solid PSF, the doughnut-shaped PSF and the subtractive PSF
Fig.6  PSFs of the confocal, the STED and the subtraction image (δ=0.5), and it can be seen the subtractive PSF is sharper than STED’s PSF [22]
Fig.7  (a) Confocal image of 20 nm fluorescence beads; (b) STED image; (c) subtraction of the confocal from the STED with δ of 0.5; (d) g-STED image; (e) subtraction image of the STED from the g-STED with ? of 0.08; (f) b′, c′, d′, e′: magnified view of the region indicated by white box in (b), (c), (d), (e)
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