3D second harmonic generation imaging tomography by multi-view excitation

Campbell, Kirby R.; Wen, Bruce; Shelton, Emily; Swader, Robert; Cox, Benjamin L.; Eliceiri, Kevin W.; Campagnola, Paul J.

doi:10.1364/optica.4.001171

Cited by 19 publications

(12 citation statements)

References 27 publications

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“…In theory close to isotropic resolution can be achieved with only 4 views but for thicker samples not all details are captured equally in every view. In a study based on SHG signal MVI, it was shown that a intertwining collagen fibre was visualized more clearly using 10 views 17 , whereas for LSFM MVI up to 24 views might be necessary to capture all the details of a large sample and to overcome local distortion due to scattering and refraction within the sample 39 . Recently it was shown that for samples not clearly visible in all views selecting pixels rich in information can provide better results than blindly incorporating all views in the MVD 39 .…”

Section: Discussionmentioning

confidence: 99%

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Multiview deconvolution approximation multiphoton microscopy of tissues and zebrafish larvae

Kapsokalyvas

Rosas

Janssen

et al. 2021

Sci Rep

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Imaging in three dimensions is necessary for thick tissues and small organisms. This is possible with tomographic optical microscopy techniques such as confocal, multiphoton and light sheet microscopy. All these techniques suffer from anisotropic resolution and limited penetration depth. In the past, Multiview microscopy—imaging the sample from different angles followed by 3D image reconstruction—was developed to address this issue for light sheet microscopy based on fluorescence signal. In this study we applied this methodology to accomplish Multiview imaging with multiphoton microscopy based on fluorescence and additionally second harmonic signal from myosin and collagen. It was shown that isotropic resolution was achieved, the entirety of the sample was visualized, and interference artifacts were suppressed allowing clear visualization of collagen fibrils and myofibrils. This method can be applied to any scanning microscopy technique without microscope modifications. It can be used for imaging tissue and whole mount small organisms such as heart tissue, and zebrafish larva in 3D, label-free or stained, with at least threefold axial resolution improvement which can be significant for the accurate quantification of small 3D structures.

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Section: Discussionmentioning

confidence: 99%

“…Despite computational power being readily available, MVI, has not found many applications in scanning microscopy, such as MPM and confocal microscopy. Indeed, some applications in MPM have been demonstrated, based on the signal of Second Harmonic Generation (SHG) from collagen fibres 16 , 17 . However, MVD has not been demonstrated.…”

Section: Introductionmentioning

confidence: 99%

Multiview deconvolution approximation multiphoton microscopy of tissues and zebrafish larvae

Kapsokalyvas

Rosas

Janssen

et al. 2021

Sci Rep

View full text Add to dashboard Cite

show abstract

“…In the case of perfect phase matching, i.e. where phase mismatch Δ = 0, emission propagates in the forward-scattered and back-scattered (FSHG, BSHG) directions 32,36,46 . Second harmonic signals, however, can be generated in other directions In the case of imperfect phase matching, where Δ ≠ 0 such as in biological tissues 43 .…”

Section: Introductionmentioning

confidence: 99%

“…Importantly, however, SHG signals are directional rather than isotropic, like in fluorescence emission (1PF or 2PF). Existing 3D imaging approaches use a variety of mechanical or computational methods 39,[46][47][48] , but all generate data through point-scanning, and use only a forward-and/or backscattered detection geometry for SHG signal collection. Since LSM uses an orthogonal signal collection configuration instead of collinear excitation and detection, it presents a frontier for SHG imaging.…”

Section: Introductionmentioning

confidence: 99%

Label-free and Multimodal Second Harmonic Generation Light Sheet Microscopy

Hanrahan

Lane

Johnson

et al. 2020

Preprint

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Light sheet microscopy (LSM) has emerged as one of most profound three dimensional (3D) imaging tools in the life sciences over the last decade. However, LSM is currently performed with fluorescence detection on one- or multi-photon excitation. Label-free LSM imaging approaches have been rather limited. Second Harmonic Generation (SHG) imaging is a label-free technique that has enabled detailed investigation of collagenous structures, including its distribution and remodelling in cancers and respiratory tissue, and how these link to disease. SHG is generally regarded as having only forward- and back-scattering components, apparently precluding the orthogonal detection geometry used in Light Sheet Microscopy. In this work we demonstrate SHG imaging on a light sheet microscope (SHG-LSM) using a rotated Airy beam configuration that demonstrates a powerful new approach to direct (without any further processing or deconvolution) 3D imaging of harmonophores such as collagen fibres in biological samples. We provide unambiguous identification of SHG signals on the LSM through its wavelength and polarisation sensitivity. In a multimodal LSM setup we demonstrate that SHG and two-photon signals can be acquired on multiple types of different biological samples. We further show that SHG-LSM is sensitive to changes in collagen synthesis within lung fibroblast 3D cell cultures. This work expands on the existing optical methods available for use with light sheet microscopy, adding a further label-free imaging technique which can be combined with other detection modalities to realise a powerful multi-modal microscope for 3D bioimaging.

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“…Despite computational power being readily available, MVI, has not found many applications in scanning microscopy, such as TPLSM and confocal microscopy. Indeed, some applications in TPLSM have been demonstrated, based on the signal of Second Harmonic Generation (SHG) from collagen bres 13,14 .…”

Section: Introductionmentioning

confidence: 99%

Multiview Deconvolution Two-photon Laser Scanning Microscopy

Kapsokalyvas

Rosas

Janssen

et al. 2020

Preprint

View full text Add to dashboard Cite

Imaging in three dimensions is necessary for thick tissues and small organisms. This is possible with tomographic optical microscopy techniques such as confocal, two-photon and light sheet microscopy. All these techniques suffer from anisotropic resolution and limited penetration depth. In the past, Multiview microscopy - imaging the sample from different angles followed by 3D image reconstruction - was developed to address this issue for light sheet microscopy based on fluorescence signal. In this study we applied this methodology to accomplish Multiview imaging with two-photon microscopy based on fluorescence and additionally second harmonic signal from myosin and collagen. It was shown that isotropic resolution was achieved, the entirety of the sample was visualized, and interference artifacts were suppressed allowing clear visualization of collagen fibrils and myofibrils. This method can be applied to any scanning microscopy technique without microscope modifications. It can be used for imaging tissue and whole mount small organisms such as heart tissue, and zebrafish larva in 3D, label-free or stained, with at least 3-fold axial resolution improvement which can be significant for the accurate quantification of small 3D structures.

show abstract

3D second harmonic generation imaging tomography by multi-view excitation

Cited by 19 publications

References 27 publications

Multiview deconvolution approximation multiphoton microscopy of tissues and zebrafish larvae

Multiview deconvolution approximation multiphoton microscopy of tissues and zebrafish larvae

Label-free and Multimodal Second Harmonic Generation Light Sheet Microscopy

Multiview Deconvolution Two-photon Laser Scanning Microscopy

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