Raman imaging through multimode sapphire fiber

We report on a sapphire fiber Raman imaging probe's use for challenging applications where access is severely restricted. Small-dimension Raman probes have been developed previously for various clinical applications because they show great capability for diagnosing disease states in bodily fluids, cells, and tissues. However, applications of these sub-millimeter diameter Raman probes were constrained by two factors: first, it is difficult to incorporate filters and focusing optics at such small scale; second, the weak Raman signal is often obscured by strong background noise from the fiber probe material, especially the most commonly used silica, which has a strong broad background noise in low wavenumbers (

High sensitivity, incoherent differential imaging

Chiara Bonati

Microscopic visualisation of optically transparent samples has been a topic of interest for several decades. Features such as density or chemical composition can influence the optical phase of transmitted light, and phase contrast can reveal these structures. Several methods of phase contrast have been developed, which can be categorised as either interferometric or non-interferometric, based on the type of coherence properties of the light used. In this work, I focus on incoherent based phase contrast, in particular Differential Phase Contrast (DPC). The choice of incoherent light brings benefits such as the absence of distortions like speckle patterns and ringing patterns, increased spatial resolution, and a simpler setup that can be used for in-vivo applications. Moreover, DPC allows reconstruction of quantitative phase maps of the samples. On the other hand, coherent-based techniques demonstrate superior performance in terms of phase sensitivity. The first part of this thesis offers a quantitative analysis of the phase sensitivity of DPC and investigates the influence of optical parameters and sample characteristics. With simulations and experiments, a relation between numerical aperture and phase sensitivity is demonstrated, and the concept of spectral matching is introduced to enhance the contrast. The methods can be generalised to any DPC setup, and allow a-priori investigation of the sensitivity of a DPC microscope at the design stage rather than through testing. Comparison between the best sensitivity that can be achieved in DPC and state-of-the-art interferometric techniques, shows that it is not possible to reach comparable single-shot performances. In this thesis, it is shown that the reason for this limitation is the strong background in DPC images, which degrades the dynamic range. DPC images are obtained with mirrored illuminations, for which the background is identical and the phase term switches in sign. The difference between these pairs of images is computed digitally, which does not improve the limited dynamic range. Lock-in DPC is proposed as a solution: instead of sampling different illumination states, lock-in DPC demodulates the phase signal when the illumination is switched, and the background is never encoded. This is enabled by periodical switching of sources coupled with a smart pixel detector, the so-called "lock-in camera". Part of this work is dedicated to the theoretical description of this method, and the analysis of the expected benefit. Experiments are also described, that demonstrate a factor of 8 improvement in single-shot sensitivity compared to standard DPC. DPC is not the only imaging technique to suffer from high intensity background: it is easy to see how the use of a lock-in camera for differential imaging can be generalised to any situation where weak modulations can be induced over a strong background. Here, an example is presented with lock-in Shifted Excitation Raman Difference Spectroscopy (SERDS). SERDS is an established Raman spectroscopy technique that takes advantage of the Raman emission spectrum being relative to the excitation wavelength to remove unwanted fluorescence emission. Two spectra with shifted excitation wavelengths are measured, their difference is computed, and the fluorescence is thus digitally removed. The parallel with DPC is immediately apparent. Both simulations and experiments are used to demonstrate the advantage of analog demodulation.

EPFL2022

Control of pulsed light propagation through multimode optical fibers

Edgar Emilio Morales Delgado

Visualization of organs and cells in the interior of living beings is a challenging task due to the light absorption and multiple light scattering occurring in biological tissue that prevents the di-rect transmission of images. A standard visualization approach is based on the use of endo-scopes which is accomplished through apertures in the body. Since their invention, the light transmission capability of optical fibers has enabled fiber optic based endoscopy with devices based on bundles of optical fibers that can directly relay images. Due to their small diameter, fiber optic endoscopes are standard in minimally invasive applications either with a white light illumination or as a fluorescent imaging device. The principle behind fluorescent endoscopy consists on the scanning of light intensity over a fluorescent sample, which is achieved by se-quential illumination of each one of the fiber bundles or by mechanical scanning of a single-mode double-cladding fiber. Collecting the fluorescent signal through the light guiding medium, an image can be reconstructed. The main drawback of fiber bundle endoscopes is the pixelated limited resolution given by the presence of thousands of cores that conform the bundle. In the double cladding fiber endoscopes, there is a limit to the miniaturization of the endoscope due to the requirement of mechanical elements at the tip of the fiber to scan the illumination over the sample. In this thesis, methods for focusing and digital scanning of optical light pulses through multimode optical fibers without any distal mechanical element are developed. Such techniques are based on spatial light modulation. Spatial phase modulation allows the control of light propagation in scrambling media, as in a multimode fiber, enabling the generation of intensity light foci or arbitrary intensity profiles. The high intensity of the transmitted pulses permits multi-photon imaging through multimode optical fibers, resulting in imaging probes of higher resolution than in the case of fiber bundles. We demonstrate in a working prototype that this approach provides all the advantages of multi-photon microscopy at the tip of an ultra-thin probe, such as a reduced photo bleaching of the sample, optical sectioning and enhanced image contrast. The developed methods can also be employed for material processing that requires high in-tensity light pulses. In specific, we demonstrate additive manufacturing, also known as 3D printing, at the tip of an ultra-thin needle. The working principle is based on two-photon polymerization, which is accomplished by scanning intense light pulses on a photosensitive material that hardens when exposed to light. All other additive manufacturing methods require very large nozzles or components in close proximity to the structure that they build, up to now. With an ultra-thin 3D printing probe we enable micro-fabrication through very small apertures, in places of difficult access or in the interior of living beings. We call it, endofabrication.

EPFL2017

Application of Adaptive Optics to Focusing and Imaging

Daniel Iwaniuk

Adaptive optics is used to abate aberrations with a wavefront correction. It is widely used in astronomy and is starting to expand into applications of laser focusing and imaging with high numerical apertures, e.g. microscopy. The physical background of focusing laser light and imaging is the same, so we can use the same methods. In both cases, light is propagating through a lens; either it becomes focused into a small region or light from a small region becomes imaged on a camera. Modulations applied on the lens to implement wavefront corrections behave similarly in both applications. A powerful simulation tool was created to characterize the impact of those modulations. As an example, we validated a design for a Fresnel lens produced on a glass fibre tip to focus its emitting light. We have developed solutions mainly for three different problems. First, a high depth of focus enables keeping a laser beam focused within a larger length or imaging objects from different positions simultaneously. In photography, this can be attained by stopping down the aperture, which introduces a huge loss of light. State-of-the-art for focusing into a line segment also shows an inefficient performance. We present an elegant lens design, which enables highly efficient elongation of the depth of focus. Preliminary studies have shown that it might be a feasible alternative for current intra-ocular lens implants in ophthalmology or for 3D visualization in imaging and microscopy. A high depth of focus also has a large potential in optical lithography and data storage, because the focal position is enlarged and does not have to be adjusted precisely to obtain a useful spot. Second, specimen-induced aberrations can affect even a perfectly adjusted, diffraction limited lens system. A planar refraction index mismatch introduces spherical aberrations, degrading the optical resolution. As a first step to correct them, we were able to characterize simultaneously the refractive index and the thickness of an unknown medium that is placed between the lens and its focal region. This was done by clever manipulation of the beam angles with the same adaptive optics element in the focusing and in the imaging system. Finally, the medium-induced spherical aberrations were corrected based on the characterization results. The point spread function degradation of a focused laser beam was completely removed, which might be useful in optical tweezers or in laser processing of biological samples. While imaging through the planar refractive index mismatch, the bending of the object field was corrected and the diffraction limited performance restored.

EPFL2012