UVG Module for ELVIS system
Make ELVIS sing—in a higher octave
This module is in addition to the near-infrared (NIR) module currently provided. This module allows for imaging of fluorescent materials which fluoresce in the ultraviolet to green (UVG) spectrum including fluorescein, coumarin-6, and related fluorophores with an excitation wavelength in the 405 nm region and an emission wavelenth greater than 500 nm.
- a UV-light LED bulb that can be threaded into any standard 120V/60W lamp socket, and
- a “green” sharp-cut filter set to thread onto the ELVIS system.
How To Use
First, set up the light source. It is ideal for this to be whatever system is most convenient to you for use in your lab. It is critical that the light system be portable and flexible enough to be brought close enough to the animal for even lighting. Imaging shown was done using a standard desk lamp.
Replace the traditional bulb with UV-LED bulb we provide. (When the light is on, do not look directly at the UV light!)
Prepare the ELVIS system and take brightfield images for alignment as described on ELVIS main page.
Note: Do NOT attach R72/NIR sharp cut filter for UVG imaging.
In order to do UVG imaging, use the 52mm UVG module provided instead of the R72/NIR sharp-cut module.
Setup and anesthetize the animal model per your institution's applicable IACUC regulations. For animals (especially those with white fur) shave the region prior to doing work, as the fur will both hinder your work and interfere with imaging. Inject or implant your desired fluorescently-labeled system. Note that the penetration capability of UVG fluorescence is not as strong as NIR fluorescence; it is best to work with relatively shallow implants (Subcutaneous; intra-muscular with small animals is fine, too, depending on the system's fluorescent brightness).
To image in UVG mode
Bring the lighted UV lamp to within 10-20 cm of the animal over the region of interest. At the same time, focus the ELVIS camera on the same region. Turn off the room lights, or put the animal in a darkened setting. Unlike NIR systems, brightly fluorescent systems in UVG mode may be visible through the viewfinder of ELVIS to the naked eye which allows easier focus and aiming.
Set the camera to manual mode. Set the F-Stop to minimum for the macro-lens and set the camera speed to 1/100. Note that unlike NIR imaging the camera will be taking pictures in roughly real-time so there is less need for strict controls on vibration. Despite this, remote shutter control is still preferred. Take a picture and check for visibility.
All example images were performed at Purdue University in accordance with Purdue Animal Care and Use Committee (PACUC) which is an OLAW approved institution. Female BALB/C mice were anesthetized with isoflurane gas and shaved in the dorsal and calf region with electric clippers. Biodegradable microparticles with a load of 10% w/w coumarin-6 fluorescent dye and ~5-10 mg/ml of particles were injected to the site via a 28 gauge syringe (100 µL SQ/50 µL IM). Imaging was done using ELVIS with a UVG module.
Q & A
Q: Can I switch back and forth from UVG to NIR modes?
A: Yes, just swap out the filters and then switch camera speed from 30s to 1/100s.
Q: How is the safety of the LED lamp relative to a traditional black light or other UV source?
A: The LED lamp is substantially safer. This lamp outputs 405 nm light almost exclusively. This avoids the significant levels of damage that can occur especially with wavelengths less than 300 nm. Additionally the LED lamp puts out significantly less heat than Hg type lamps and can be turned on/off rapidly without warm-up.
Q: But I already have an Hg black-light (or alternate UV source) set up in my lab and ready to go. Will the ELVIS module work with that?
A: Yes, you can also use any source which will excite your fluorophore.
Q: What is the tissue penetration of this relative to NIR?
A: Substantially less, you will need to work with shallow systems for UVG.
Q: Why does the “fur” glow, is it fluorescent in UV-G region?
A: Unfortunately even with the best sharp-cut filter some reflected light still passes through. Due to this white colored (or highly reflective) objects will seem to glow fluorescently. It is best to shave the region of the animal that you desire to image as well as use a black background for imaging.