Frequently Asked Questions
Luciferase seems more convenient. Why and When should I use NIS?
Luciferase is an excellent reporter gene for ‘quick and dirty’ studies of cell or virus fate in SCID or nude mice. Up to five mice can be imaged at one time making luciferase an excellent screening tool, and small numbers of cells can be detected, but images are two-dimensional and resolution is poor.
Importantly, researchers from the University of Minnesota recently reported that firefly luciferase is immunogenic, resulting in substantial loss in gene expression levels in immunocompetent mice over time. Hence, luciferase is not desirable for long term cell tracking studies.
Reference: Cellular immune response against firefly luciferase after sleeping beauty-mediated gene transfer in vivo. Podetz-Pedersen KM, Vezys V, Somia NV, Russell SJ, McIvor RS. Hum Gene Ther. 2014 Nov;25(11):955-65. doi: 10.1089/hum.2014.048. Epub 2014 Sep 22. PMID:25093708
Which reporter gene should I use for whole animal imaging?
Mice: you can use optical imaging (luciferase or fluorescent proteins) or nuclear imaging (NIS). Biolumonescence imaging is very sensitive and there is no issue with background signals. Due to issues with autofluorescence from fur or rodent chow, fluorescence imaging in living animals can be challenging as it requires you to first subtract the background fluorescence signals. If you want to obtain high resolution 3D true tomographic data, you should use nuclear reporter genes (e.g. NIS) with SPECT/CT or PET/CT imaging for best quantitative results.
Rats or larger: Due to inability of light to penetrate deep tissues and scatter, nuclear reporter genes (e.g. NIS) are the ideal genes for in-life imaging of living animals. If you need to monitor the genes long term, you will need to ensure that you use an immunogenic reporter gene (e.g. species specific NIS) to prevent host immune mediated rejection of NIS transduced cells.
I live outside the USA. Do I need an import license to receive the viral products?Each country has its own set of rules and regulation regarding import of infectious agents. Shipping will be on dry ice via Fedex courier service. A valid import permit might be needed before the product can be shipped to the designated country. We will work with you to ensure safe and timely delivery of the products ordered. Do not worry! We have successfully shipped our products internationally and within the United States.
Do I need biosafety approval to receive OVs?
Imanis will be able to ship the OVs to you after you have ordered them. However, most institutes require the scientist and lab to have biosafety registration and approval from the institutional biosafety review committee (IBC) prior to using and experimenting with any recombinant viruses. Please treat all oncolytic viruses as a biohazard and take the necessary actions to ensure personnel and environment protection.
For labs in the USA requesting VSV viruses, you will need to obtain a license from the United States Department of Agriculture (USDA) to certify that your lab and personnel are qualified to handle and transport VSV agents. Please refer to the USDA website for more information. NOTE: The VSV offered are derived from cDNA encoding attenuated lab adapted Indian strain of VSV and NOT wild type VSV.
Can I add my viruses to the repository?
Yes you can! We would love to offer your virus for research use!
Please contact us at firstname.lastname@example.org or call us at +1-507-218-2559 (8:30 am to 5:00 pm, USA Central Time zone) to discuss further.
What if I need an OV or lentivector not in the product list?We can custom design and make it for you! Please contact Imanis at email@example.com or call us at +1-507-218-2559 (8:30 am to 5:00 pm USA Central Time zone) to discuss your research needs.
Can I grow the OVs and propagate new stocks in my lab?Yes! You can purchase the 'standard titer' stocks for this application (0.25 mL,
about 106 TCID50/mL). Each product will be shipped on dry ice and comes with a certificate of analysis and a protocol for amplification of new viral stocks in your lab.
Can the OVs be administered to animals intravenously?
Yes! The Ovs are suspended in biocompatible media and can be injected intravenously into animals. We suggest dilution of viruses to desired dose level in sterile normal saline and kept on ice until use.
The high titer and high purity viral stocks (listed as High Titer) are prepared specially for in vivo antitumor efficacy experiments. Many of our clients (including our research team) find these high titer stocks very appealing for studies in mice.
Are the OVs well characterized and accurately titered?Yes! Each of the production runs are fully characterized by Imanis scientists and have to pass QC tests. The product will come with its certificate of analysis and are ready to use immediately in your experiments.
Which viruses are offered?Imanis has exclusive rights to offer replication competent recombinant measles viruses and vesicular stomatitis viruses. Many of these viruses encode reporter genes, including NIS, to meet for your research needs. Vaccinia and herpes simplex viruses are coming soon!
Can NIS be used for long term studies?Yes. Unlike luciferase, which is highly immunogenic in mammals, NIS does not provoke an immune rejection response and does not affect the survival or biology of the NIS-marked cells which can therefore be studied throughout the life of the experimental subject.
How many times can I NIS image a single subject?In principle, a single animal can be imaged numerous times. But because animals have to be anesthetized for each imaging session, the number of images that can be obtained is limited by IACUC rules governing exposure to anesthetic agents. In human subjects anesthesia is not required so the number of imaging studies is limited by international radiation exposure standards.
How often can I repeat the NIS imaging study in the same subject?This depends on the physical half life of the radiotracer you are using. Basically, you need to be sure that your imaging study is not contaminated by residual radioactive signals from the preceding study. The SPECT tracer 99mTcO4 (pertechnetate) has a half life of 6 hours so imaging can be repeated daily. The PET tracer B18F4 (tetrafluoroborate) has a half life of only 110 minutes so imaging can be repeated several times in a day. If you have questions about a specific radiotracer, email to firstname.lastname@example.org
What is a typical imaging protocol to detect NIS positive cells in a mouse?Imaging of a mouse previously implanted with NIS-cells or injected with NIS-vectors.
Mouse is given 250-300 uCi of SPECT or PET isotopes intravenously (tail vein) or intraperitoneally.
- Wait 1-2 hours.
- Place anesthetized mouse on the imaging bed.
- Image per protocol
Is NIS safe to use?The gene itself is very safe to use, but keep in mind that the lentivectors containing the gene are rated Biosafety Level 2 (BSL2) and radiation training is generally required for researchers who will be conducting the 1-125 uptake assay using out NIS-IT kits. For those wanting to avoid using radiation in the laboratory, we do offer a NIS-YFP vector that allows for a non-radioactive uptake assay to analyze NIS function.
How can NIS technology help me?If you want to study dynamic processes in living animals, such as the migration, proliferation and death of specific cell populations, changes in gene expression, or the spread of virus infections, then NIS is most probably the ideal reporter gene to serve your needs.
What is NIS?NIS is a snitch. It transports radioactive iodide and other ‘NIS radiotracers’ into genetically marked, NIS-expressing cells which can then be ‘seen’ by noninvasive gamma camera, SPECT or PET imaging such that their location in the body can be accurately pinpointed.
What are the acceptable formats for the image analysis files?We accept most data files from the imaging machines. DICOM and NIFITI files are the most common files that are received by our analysts. If necessary, data files can be re-loaded and save into the preferred files formats.
I need help with analyses of my image files!We can help! Please email Imanis to enquire. We will first have a consultation with you to determine your needs, review your data files to determine the best course of action to get the most out of your data. We can provide ROI (region of interest) analysis, provide you with presentation or publication ready images and movies. Let's talk!