Planar Airy beam light-sheet for two-photon microscopy

We demonstrate the first planar Airy light-sheet microscope. Fluorescence light-sheet microscopy has become the method of choice to study large biological samples with cellular or sub-cellular resolution. The propagation-invariant Airy beam enables a ten-fold increase in field-of-view with single-photon excitation; however, the characteristic asymmetry of the light-sheet limits its potential for multi-photon excitation. Here we show how a planar light-sheet can be formed from the curved propagation-invariant Airy beam. The resulting symmetric light sheet excites two-photon fluorescence uniformly across an extended field-of-view without the need for deconvolution. We demonstrate the method for rapid two-photon imaging of large volumes of neuronal tissue.Comment: 7 pages, 4 figure

Terahertz-imaging with demodulating detector array

Diese Dissertation beschäftigt sich mit Bildgebung im Terahertz-Frequenz-bereich, der im Spektrum der elektromagnetischen Strahlung zwischen den Radiowellen und dem Licht angesiedelt ist. Der elektrooptische Detektionsmechanismus ist besonders dazu geeignet, einen großen Frequenzbereich (rund 4 Oktaven) und gleichzeitig eine Vielzahl von Messpunkten in nur einer Messung abzudecken. Die Spektral- und Bildinformation wird dabei in einem Detektorkristall vom Terahertz-Strahl auf einen optischen, nahinfraroten Laserstrahl übertragen und dann in eine Intensitätsmodulation desselben übersetzt. Um die sehr gering ausfallende Modulationstiefe erfolgreich vom unerwünschten Hintergrund zu trennen, wird die PMD-Kamera (Photonic Mixing Device) als demodulierendes Detektorarray eingesetzt. Sie bestimmt dabei ortsaufgelöst die Amplitude der Intensitätsmodulation, von der sich auf die elektrische Feldstärke des THz-Pulses im Detektorkristall zu einem sub-ps Zeitpunkt schließen lässt. Durch einen Scan im Zeitbereich kann so der gesamte Terahertz-Puls an 3072 Orten parallel rekonstruiert werden, wodurch Bildgebung im sub-ps-Zeitbereich und im Terahertz-Frequenzbereich ermöglicht wird. Nach diesem Prinzip sind zwei verschiedene Aufbauten realisiert und eingehend charakterisiert worden. Beispielmessungen belegen, dass nun Multipixel-Terahertz-Bildgebung auch ohne die Notwendigkeit von Verstärkerlasern möglich ist und machen die erreichte Optimierung deutlich.This dissertation is about imaging in the domain of terahertz-frequencies which is located between radio waves and light in the electromagnetic spectrum. Electrooptic detection is especially convenient to cover a wide spectrum (approximately 4 octaves) while at the same time enabling for parallel measurements at many points. Using a detector crystal, the spectral and spatial information in the terahertz beam is being transferred to the (optical) laser beam and then converted into an intensity modulation. At this point the PMD-Camera (Photonic Mixing Device) is used as a demodulating sensor array to seperate the resulting very small intensity modulation successfully from the unwanted huge background. It determines laterally resolved the amplitude of the intensity modulation, from which the electrical field magnitude of the terahertz pulse in the crystal at a sub-picosecond instant of time can be derived. While scanning in time domain the whole THz pulse can be reconstructed for 3072 different pixels in parallel enabling imaging with sub-picosecond resolution in time domain and in the region of THz frequencies. Two different imaging setups using this scheme were realized and exhaustively characterized. Exemplary measurements show that multipixel terahertz imaging now is possible without the need for an amplified pulse laser system and they illustrate the achieved optimization

Label-free and multimodal second harmonic generation light sheet microscopy

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 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