In the last three decades, optical trapping techniques were heavily employed for contactless trapping and manipulation of biological samples. Dual-beam laser traps (DBLT) proved their convenience and became widely used as biophysical tool once a simplified experimental setup was proposed. This simplification was achieved by replacing the two objectives with optical fibers to deliver the two counter-propagating laser beams. However, fiber alignment can be inconvenient, time consuming and requires a lot of practice. Here, we present a novel way to overcome these issues by combining reconfigurable diffractive optical elements (DOE) and two photon lithography (2PL), using a single low NA objective. A single laser beam is divided into several beams by displaying a DOE on a spatial light modulator (SLM). This allows us to dynamically reconfigure the number of the beams, their shape, and relative 3D alignment. Furthermore, we use 3D printed micro-mirrors to direct the laser beams against each other and obtain a DBLT. The micro-mirrors were fabricated on top of a coverslip, by means of 2PL. Our preliminary results show the ability to trap dielectric and biological samples and their full 3D manipulation in a DBLT configuration. The ability to use DOEs to set the number of beams and their shape allow this technique to be coupled with novel forms of microscopy.
The viscoelastic material properties of biological systems are increasingly recognized as important parts of signaling cascades involved in developmental and pathological processes. They are furthermore assumed to play a crucial role in surviving extreme environmental conditions for certain organisms, such as yeast cells. Confocal Brillouin microscopy gives access to the viscoelastic material properties of single cells and tissues in a contact- and label-free manner and with a high spatial resolution. In combination with quantitative phase imaging, it is then possible to determine the longitudinal modulus and the viscosity of the sample. In this study, we probed living zebrafish larvae in all anatomical planes, at different time points during development and after spinal cord injury. We could show, that confocal Brillouin microscopy detects the viscoelasticity of different anatomical structures without affecting the animal’s development. We furthermore observed a transiently decreasing Brillouin shift after spinal cord injury and a difference in Brillouin shift between in vivo and ex vivo measurements of the same sample region. Using quantitative phase imaging we additionally show, that the Brillouin shift of the probed tissues is mainly governed by their longitudinal modulus and viscosity. In conclusion, this work constitutes the methodical basis to identify key determinants of viscoelastic tissue properties during biologically important processes in vivo.