Beyond the standard imaging techniques of brightfield and fluorescence microscopy, the Microscopy SRF offers a wide variety of other imaging capabilities for your research needs. Past experiments have included:
- Second Harmonic Generation Imaging
- Dye Separation
- Extended Focus Imaging
- Tile Scanning
- Plane Polarized Light Imaging
- Circularly Polarized Light Imaging
Don’t see an application you need here? Contact us today to set up a free consultation to discuss your imaging needs!
Second Harmonic Generation
Looking for a solution to image collagen without the need for immunostaining? Second Harmonic Generation (SHG) imaging might just meet your imaging needs. SHG is an optical phenomenon produced by certain biological materials with ordered structural protein assemblies such as collagen fibers. When photons of a given frequency interact with these materials, they generate new photons with twice the energy and half the wavelength. For example, collagen imaged at 850nm will emit light at 425nm. Samples such as these can be imaged on the Olympus FV1000 MPE multiphoton microscope without the use of fluorescent proteins or antibody stains.
Tendon model of damage induction; collagen imaged by SHG. Imaged using Olympus FV1000 multiphoton microscope. Courtesy of Stephen Ros, Evan Flatow Lab.
Seeing bleed-through (or crosstalk) in your images? Using emission spectra as references, combined or mixed emission signals can be clearly separated into the dyes that make up the total signal. This function is available on all the Microscopy SRF’s laser scanning confocal systems:
- LeicaSP5-DM confocal
- LeicaSP5-DMI confocal
- LSM510 confocal (separation known as “Linear Unmixing”)
Mouse kidney section stained with Alexa Fluor 568 phalloidin (red), Alexa Fluor 488 WGA (green), and DAPI-nuclei (blue). Original image was acquired using LeicaSP5-DM confocal (left) and dye separation of these signals was performed within the LAS AF software (right). Sample courtesy of Molecular Probes (FluoCells Prepared slide #3).
Extended Focus Imaging
Trying to image a large sample and having trouble keeping it in focus? The extended focus imaging capability can resolve this issue by extending the focus beyond the limited depth of focus of brightfield microscopes. This tool combines images acquired at many different focus points and extracts sharply imaged details to provide a single image that is entirely in focus. The Extended Focus module is available on the Axioplan2IE and the Olympus MVX10 Stereoscope.
Drosophila imaged on the Axioplan2IE. Single image acquired within the Z-series (left) and Extended Focus image (right). Sample courtesy of Sujin Bao.
For larger samples that require high resolution imaging, we offer systems that are capable of tile scanning and stitching a mosaic of images. Whole tissue slices or other large areas can be acquired using automated motorized stages available on a variety of modalities, including epi-fluorescence, brightfield, confocal, and multiphoton. Microscopes capable of tiling include:
- Axioplan2IE (MosiaX)
- DM6000 (Tile Scan)
- Olympus MVX10 Stereoscope (Manual Tile Acquire/Arrange)
- Olympus IX70 LiveCell (Autoscan: Ox Plowing)
- LeicaSP5-DM confocal (Tile Scan)
- LeicaSP5-DMI confocal (Tile Scan)
- Olympus FV1000 MPE multiphoton (Mosaic)
Mouse brain montage made up of 80 individual images acquired using the 10x/0.5 NA objective and motorized stage installed on the Axioplan2IE. Image courtesy of Jing Wang, Zhenyu Yue lab.
Plane & Circularly Polarized Light Imaging
Polarized light microscopy can be used to image birefringent biological samples. The Axioplan2 system is capable of both plane and circularly polarized light microscopy.
Lamb hoof stained with picrosirius red, imaged using brightfield (A), plane polarized (B), and circularly polarized (C) light microscopy. Sample courtesy of Virginia Gillespie.
Demonstrations and Seminars
Interested in a technology not currently offered at the Microscopy SRF? Contact us regarding your imaging or analysis needs!
The Microscopy SRF hosts educational microscopy workshops that give users the hands-on opportunity to learn about and demo new microscope systems. The most recent workshop involved a demo of the Zeiss ELYRA superresolution microscope with SIM and PALM/dSTORM capabilities. Users were assisted by Zeiss applications specialists and were able to acquire both confocal and superresolution images for comparison.
Huh-7.5 cells replicating a chimeric hepatitis C virus (HCV, APP144) imaged on the Zeiss LSM710 confocal (left) and ELYRA superresolution microscope system (right). The HCV core protein was detected using two primary mouse monoclonal antibodies (gift from Abbott Labs) and secondary anti-mouse antibodies conjugated to Alexa Fluor 488 (green) and Alexa Fluor 633 (red). Nuclei labeled with DAPI (blue). Courtesy of Erin Doyle, Andrea Branch Lab.
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