Motivation for the study
Tissue engineering is an evolving field of medicine that aims to grow new viable tissue for repairing or regenerating damaged tissues in the body. These tissue constructs are typically made by placing cells in support matrices that have proper growth-inducing factors. However, visualizing the development and composition of these tissue constructs is challenging with currently available optical imaging methods: they are either not capable of producing 3D images at sufficient depth or lack the bright-field imaging functionality. To answer this need, a high-resolution 3D imaging microscope was designed and constructed by combining two different imaging methods into one system. This setup combines advantages of both methods and provides more information from the sample. The system was then tested by imaging two different biological samples.
Multimodal imaging setup
The imaging system was built by the Computational Biophysics and Imaging research group. Optical projection tomography (OPT) and selective plane illumination microscopy (SPIM) were combined into one multimodal imaging system. SPIM is fluorescence-based imaging method that projects a single sheet of light to illuminate only a thin area of the sample that is being imaged. This single frame imaging significantly decreases photobleaching and enhances imaging speed compared to standard confocal microscopy that relies on scanning mechanism. However, it can only visualize fluorescent parts of the sample, so OPT was combined with SPIM to create context to images by enabling simultaneous bright field imaging of the sample.
The microscopy system was set up horizontally. This arrangement allows having the exact same detection path for both imaging methods, since SPIM illuminates the sample from the side while OPT illuminates parallel to the detection path. LED provides illumination for OPT while SPIM has illumination from Modulight’s ML6600 laser with three wavelengths (488, 561, 638 nm) for efficient excitation of fluorophores. Laser was specifically designed to be stable even at very low powers (<40 mW) to enable longer periods of continuous imaging without fast photobleaching and phototoxicity to the sample.
Imaging biological samples with the system
The multimodal imaging system was used to study the structure of two biological samples: fish head vasculature and nuclear lamina. Samples were imaged across the z-axis by taking an image at regular intervals. After imaging, a 3D model of the sample was created. Fluorescent labels (green fluorescence protein expressed in fish endothelial cells, or Alexa 561 immunostained nuclear lamina) were excited with integrated lasers to generate fluorescence signal for SPIM imaging.
Fish head vasculature
Videos courtesy of T. Montonen / Computational Biophysics and Imaging research group / TUNI
Related Modulight products and Services
Bioactive glass ions induce efficient osteogenic differentiation of human adipose stem cells encapsulated in gellan gum and collagen type I hydrogels
Kaisa Vuornos, Miina Ojansivu, Janne T. Koivisto, Heikki Häkkänen, Birhanu Belay, Toni Montonen, Heini Huhtala, Minna Kääriäinen, Leena Hupa, Minna Kellomäki, Jari Hyttinen, Janne A. Ihalainen, Susanna Miettinen
Materials Science and Engineering: C, 2019, 99 (905-918)
Multimodal Optical 3D-Imaging System – Hardware and Software
Master’s Thesis, 2017
Do you have questions or comments related to this application note? Maybe you would like to request literature? Or would you like us to feature your research? Please drop us a line!