In Vivo Imaging System - Newton 7.0, Vilber
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The Newton 7.0 is a highly sensitive optical imaging system dedicated to the visualization of in vivo, in vitro and ex vivo applications.
Applications in brief
Various bioluminescent reporters like firefly luciferase and many fluorescent molecular reagents can be used to visualize and track tumor, disease or inflammation development, target molecules to nanoparticles or follow biodistribution and pharmacokinetics studies within living aninals in a non-invasive manner.
Features
- Detection of bioluminescence and multispectral fluorescence
- Timelapse acquisition for real-time distribution studies
- 3D Bioluminescence Tomography (BLT) enhancing traditional planar 2D bioluminescence imaging into quantifiable three-dimensional output
- Co-registration with digital Organ & Bones atlas for digital anatomy reconstruction of the imaged animal in various postures
Which samples may I use?
Selected samples, but not limited to:
- live anesthetized mice (up to 5 simultaneously)
- live anesthetized rats (up to 3 simultaneously)
- dissected organs placed in e.g. standard petri dishes
Location
The Newton is placed in the Skou building animal facility. Therefore, please familiarize yourself with guidelines and regulations regarding animals and samples for imaging.
Training and use
Please note: The Newton Software and Hardware Manual applies to a previous software version, hence there are some discrepancies.
Tips and tricks
Bioluminescence or fluorescence?
- Bioluminescence imaging is based on endogenous production of light by expression of the enzyme luciferase upon reacting with the substrate luciferin in the presence of oxygen and ATP. Since bioluminescent imaging does not require an excitation light source, the level of background (often caused by autofluorescence) is extremely low. Hence, bioluminescence imaging enables imaging of processes that produce minimal signal, e.g. lymphocyte trafficking or detection of small numbers of cancer cells.
- Fluorescence results from a process that occurs in molecules known as fluorophores. Typically, acquisition of a fluorophore signal relies on excitation by an external light source. The fluorophore absorbs the excitation light, reaching a higher energy state. By returning to its former state, it emits fluorescent light. Fluorescence imaging is widely used for e.g. gene expression and protein detection.
Which fluorophores?
- Consult our Specifications section for filter bandwiths.
- In case of multiple fluorophores, consult a spectra viewer to estimate the risk of cross-talk/bleed-through.
Consider the eyes
- Following anaesthesia, ensure to protect the eyes of your animals from drying out by applying eye ointment with a clean cotton wool bud.
Did my experiment work at all?
- Obtain a quick image for swift evaluation by adjusting the binning settings.
Consider autofluorescence
- Use a control mouse to investigate the degree of autofluorescence.
- Consider alfalfa-free chow for 4-7 days prior to imaging to reduce autofluorescence caused by phytochromes in the abdominal region.
- The fur of mice may mask your results particularly when detecting fluorescence in the blue-green spectrum. Consider to remove the fur by shaving and/or depilatory cream and/or to work in the red spectrum.
- The tail of mice is highly autofluorescent. Consider to cover the tail with e.g. black paper (located on site in drawer under Newton) prior to imaging.
- In case of imaging in the abdominal region, consider emptying the bladder prior to imaging. Contact stable personnel for that matter.
- In case of strong fluorescence, consider using spacers between each animal. Spacers are located inside the door of the Newton 7.0.
Location of area of interest
- For imaging under bone structures such as the brain and spinal cord, choose a bioluminescence setup rather than fluorescence as these areas are difficult to illuminate sufficiently.
- For deeply located areas, choose colours in the red spectrum (650-900 nm).
Spectral unmixing
- Your fluorophores may have a significant overlap of excitation and emission spectra, also known as crosstalk or bleed-through. With SpectraViewer, you can determine overlap of your fluorophores. In case of overlap, you may have to consider spectral unmixing to be able to distinguish between the signals. Spectral unmixing can only be done on-site while acquiring the image.
- Read more about spectral unmixing
- If you need further spectal unmixing files, please contact the Bioimaging Core Facility for guidance.
Specifications
| Full equipment name | Newton 7.0 FT500, Vilber |
| Camera | DarQ-9 1"Scientific CCD Sensor, 2160x2160 (4.6 MP) |
| Lens | f/0.70 |
| Fluorescence spectrum | 400>900nm |
| Fluorescent channels | 420 / 480 / 520 / 580 / 640 / 680 / 740 / 780 nm |
Filter bandwidths (narrow band-pass emission filters) | F-500 (530-550nm), F-550 (550-580nm), F-600 (580-640nm), F-650 (640-670nm), F-700 (690-720nm), F-750 (710-760nm), F-800 (800-840nm), F-850 (830-870nm) |
| Bioluminescence | yes |
| Field of view | 6 x 6 cm to 20 x 20 cm |
| Animal capacity | up to 5 mice or 3 rats simultaneously |
| Heated stage | yes, adjustable temperature |
FAQ
Animal Facility
- How do I get access to the Skou animal facility?
- In order to access the Skou animal facility, you will need to book a training session with the stable personnel first. Please directly contact the stable personnel for this matter.
- Which samples may I bring to the stables?
- You are NOT allowed to bring animals from other facilities to the Skou animal facility.
- Please contact the stable personnel for information on which and how to bring samples to the Skou animal facility.
Equipment
- How should I book the Newton7.0?
- To ensure that you have enough space in the room, in which the Newton 7.0 is located, please always book the Newton 7.0 as well as the room itself.
- The Newton 7.0 can be booked via the Bioimaging Core Facility booking system
- The room can be booked via the outlook system and is called '1115-K18A IVIS' in the outlook booking system. Add the room in the 'invite person' field. In case of doubts on how to book the room, contact IT Support.
- Which isoflurane settings should I use?
- Follow isoflurane anesthesia-guides located next to the isoflurane equipment.
- The oxygen carrier (white container on wheels on the floor) adjusts its flow automatically according to your isoflurane settings. Please do not adjust oxygen carrier setting.
- Isoflurane vaporizor (adjustable by turning the wheel/top): 4-5% for induction chamber, 2,5-3% for maintenance in nose cones
- Isoflurane induction flow (for initial anesthesia in induction chamber): 1 L/min
- Isoflurane flowmeter (labelled 'manifolder', i.e. supplies the Newton with isoflurane): 1-1.5 L/min. The isoflurane flowmeter does not need to be adjusted according to number of nose cones in use.
- Isoflurane settings may need to be adjusted according to particular species or strains of animals.
- My animal keeps waking up inside the Newton, what can I do?
- Follow isoflurane anesthesia-guides located next to the isoflurane equipment.
- Isoflurane settings may need to be adjusted according to particular species or strains of animals.
- Ensure that the animal is sufficiently anesthetised in the induction chamber prior to transfer to the nose cone. This may take 2-5 min.
- Ensure that the nose of the animal is placed as close as possible to the isoflurane entry site inside the nose cone (at the very back of the nose cone). Even a small distance to the isoflurane entry site results in much lower isoflurane amounts.
- Consider turning on the isoflurane flow towards the Newton half a minute prior to moving the animal from the induction chamber to the nose cone, ensuring that isoflurane has already reached the nose cone prior to moving the animal. No need to adjust vaporiser setting for this.
- How should I clean the equipment after use?
- After use, please clean the instrument (stage and nose cones) by wiping with water and ethanol on a tissue. Never use Virkon (pink fluid) as it damages the surface of the instrument and increases background fluorescence.
- Do not spray liquids inside the instrument as this may damage light sources, electrical components and the camera lens.
- Please leave the work station neat and clean for the next user.
- How shall I leave the isoflurane equipment?
Follow the step-by-step guide next to the isoflurane equipment.
Please make sure to:
- refill the isoflurane container
- weigh the isoflurane scavengers
- write the weight of the isoflurane scavenger on the scavenger lable
- ensure to contact the stable personnel if the scavenger has to be replaced
- Is isoflurane included in the price?
- Isoflurane is not included in the price for booking of the Newton 7.0 via the Bioimaging Core Facility. Please contact the stable personnel for guidance on obtaining isoflurane.
- I am pregnant or planning a pregnancy. Is it safe to use the Newton 7.0?
- As isoflurane is harmful for the foetus, please ensure that you comply to all safety measures needed. If in doubt, contact the stable personnel for guidance and protective equipment.
Data Storage
- How can I get access to my data?
- After using a microscope or a workstation, please transfer your data immediately to your personal AU drive (O:) and clean up your local directories on instrument PCs if you are not reusing data in subsequent experiments.
- The Bioimaging Core Facility staff will provide support to non-AU users for exporting data.
- Use of USB memory sticks or external hard drives is for security reasons not allowed.
- Does the Bioimaging Core Facility regularly back up the user data?
The Bioimaging Core Faility is not responsible for your data. DATA ON LOCAL PC DRIVES WILL BE DELETED ON A REGULAR BASIS in order to keep the systems performing optimally.
Please ensure that you transfer your data to your personal AU drive (O:) after each session.
- May I use my personal USB memory stick or external hard drive for data storage?
No, any use of USB memory sticks or external hard drives is for security reasons not allowed. Usage of USB memory sticks may lead to suspension of the user license.
Data Analysis
- How may I process and analyse my data?
- Obtained images may be processed and analysed using the freeware Kuant. The freeware can be downloaded upon registration. Contact us for the system's serial number.
- Kuant is compatible with Windows.
- If need be, we have installed Kuant on our workstation PC3. Please ensure to book your time slot in our booking system.
Publications
- How shall I refer to the equipment?
- The purchase of the Newton 7.0 has been possible by a grant from the Carlsberg Foundation. Please acknowledge the Carlsberg Foundation (CF23-1388) in publications.
- Please also ensure to acknowledge the Bioimaging Core Facility. For guidance consult Publications & Acknowledgements.
Need help?
Come to our Open Office sessions to discuss your imaging needs or get advice on workflows, free of charge: