All-optical microscope autofocus based on an electrically tunable lens and a totally internally reflected IR laser

Funding Information: Deutsche Forschungsgemeinschaft (SFB 688); NIH (Prime Grant No. 5 R01 DA038882-02); University of Wuerzburg.Microscopic imaging at high spatial-temporal resolution over long time scales (minutes to hours) requires rapid and precise stabilization of the microscope focus. Conventional and commercial autofocus systems are largely based on piezoelectric stages or mechanical objective actuators. Objective to sample distance is either measured by image analysis approaches or by hardware modules measuring the intensity of reflected infrared light. We propose here a truly all-optical microscope autofocus taking advantage of an electrically tunable lens and a totally internally reflected infrared probe beam. We implement a feedback-loop based on the lateral position of a totally internally reflected infrared laser on a quadrant photodetector, as an indicator of the relative defocus. We show here how to treat the combined contributions due to mechanical defocus and deformation of the tunable lens. As a result, the sample can be kept in focus without any mechanical movement, at rates up to hundreds of Hertz. The device requires only reflective optics and can be implemented at a fraction of the cost required for a comparable piezo-based actuator.Publisher PDFPeer reviewe

High resolution autofocus for spatial temporal biomedical research

published_or_final_versio

Development of a handheld fiber-optic probe-based raman imaging instrumentation: raman chemlighter

Raman systems based on handheld fiber-optic probes offer advantages in terms of smaller sizes and easier access to the measurement sites, which are favorable for biomedical and clinical applications in the complex environment. However, there are several common drawbacks of applying probes for many applications: (1) The fixed working distance requires the user to maintain a certain working distance to acquire higher Raman signals; (2) The single-point-measurement ability restricts realizing a mapping or scanning procedure; (3) Lack of real-time data processing and a straightforward co-registering method to link the Raman information with the respective measurement position. The thesis proposed and experimentally demonstrated various approaches to overcome these drawbacks. A handheld fiber-optic Raman probe with an autofocus unit was presented to overcome the problem arising from using fixed-focus lenses, by using a liquid lens as the objective lens, which allows dynamical adjustment of the focal length of the probe. An implementation of a computer vision-based positional tracking to co-register the regular Raman spectroscopic measurements with the spatial location enables fast recording of a Raman image from a large tissue sample by combining positional tracking of the laser spot through brightfield images. The visualization of the Raman image has been extended to augmented and mixed reality and combined with a 3D reconstruction method and projector-based visualization to offer an intuitive and easily understandable way of presenting the Raman image. All these advances are substantial and highly beneficial to further drive the clinical translation of Raman spectroscopy as potential image-guided instrumentation

Nonmechanical parfocal and autofocus features based on wave propagation distribution in lensfree holographic microscopy

Performing long-term cell observations is a non-trivial task for conventional optical microscopy, since it is usually not compatible with environments of an incubator and its temperature and humidity requirements. Lensless holographic microscopy, being entirely based on semiconductor chips without lenses and without any moving parts, has proven to be a very interesting alternative to conventional microscopy. Here, we report on the integration of a computational parfocal feature, which operates based on wave propagation distribution analysis, to perform a fast autofocusing process. This unique non-mechanical focusing approach was implemented to keep the imaged object staying in-focus during continuous long-term and real-time recordings. A light-emitting diode (LED) combined with pinhole setup was used to realize a point light source, leading to a resolution down to 2.76 μm. Our approach delivers not only in-focus sharp images of dynamic cells, but also three-dimensional (3D) information on their (x, y, z)-positions. System reliability tests were conducted inside a sealed incubator to monitor cultures of three different biological living cells (i.e., MIN6, neuroblastoma (SH-SY5Y), and Prorocentrum minimum). Altogether, this autofocusing framework enables new opportunities for highly integrated microscopic imaging and dynamic tracking of moving objects in harsh environments with large sample areas

Design and implementation of the SBX1: a smart environment chamber for biological research and discovery

2021 Summer.Includes bibliographical references.Modern biomedical laboratories make significant use of environmentally controlled chambers for incubation and examination of live cell samples. They require precise control over temperature, humidity, and gas concentration to mimic natural conditions for cell survival and growth. Many incubators and live cell imaging systems exist as commercial products; however, they are prohibitively expensive, costing tens or hundreds of thousands of dollars depending on capabilities of the system. This thesis presents the electrical, optical, mechanical, and software design of the SBX1Smart Environment Chamber. This device aims to fulfill the needs of most users at a lower cost than current commercial offerings, providing an opportunity for less funded labs to pursue biomedical research and development. The chamber provides temperature, humidity, and gas concentration controls, an internal microscope with an automated stage, and an integrated ARM microcomputer to with a graphical user interface for control and monitoring of the system. A patent has been filed for the SBX1; application no. US 2020/0324289 A1

Mueller Matrix Microscopy for In Vivo Scar Tissue Diagnostics and Treatment Evaluation

Scars usually do not show strong contrast under standard skin examination relying on dermoscopes. They usually develop after skin injury when the body repairs the damaged tissue. In general, scars cause multiple types of distress such as movement restrictions, pain, itchiness and the psychological impact of the associated cosmetic disfigurement with no universally successful treatment option available at the moment. Scar treatment has significant economic impact as well. Mueller matrix polarimetry with integrated autofocus and automatic data registration can potentially improve scar assessment by the dermatologist and help to make the evaluation of the treatment outcome objective. Polarimetry can provide new physical parameters for an objective treatment evaluation. We show that Mueller matrix polarimetry can enable strong contrast for in vivo scar imaging. Additionally, our results indicate that the polarization stain images obtained form there could be a useful tool for dermatology. Furthermore, we demonstrate that polarimetry can be used to monitor wound healing, which may help prevent scarring altogether