In Vivo Imaging - Science method
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Questions related to In Vivo Imaging
I am interested in the sensory innervation of the zebrafish caudal fin between 8 and 30 dpf. I am planning to repeatedly image the tail fin during this time span using a confocal microscope. Since it is in vivo imaging, there is the issue of preventing pigmentation (albino lines are unfortunately not an option). Does anyone have experience using PTU after 5 dpf (that is the only time point until I used it for now) or has any other method/ideas to prevent/minimize pigmentation of the fin until 30 dpf?
Thanks a lot!
I would like to know if it is possible to use the the calcium indicator dyes to stain tissues in a live animal.
I am not talking about genetically encoded calcium indicators of course. My main interest for this question is common calcium dyes such as Flu 4, etc.
What is In Vivo Microscopy (IVM) images?
I'm not clear about this term, can anyone please point out it?
Any ideas and resource links are much appreciated🖤
I am following a protocol from Ana G Barata's (who is no longer in research and has no contact info):
Basically, the goal is to measure ratiometric changes in roGFP2 fluorescence (405/488 nm excitation).
Because 405 excitation gives noisy images they recommend thresholding aside from routine background removal.
My specific issue is that thresholding creates binary images (without brightness gradients), so the ratios (using the image calculator as in Barata’s and Morgan’s papers) are also binary.
I'd appreciate any specific advice or if someone has a protocol to share.
I would like to know if any carbon allotrope based nanomaterials (such as fullerenes, nanotubes, graphene, reduced graphene, graphene oxide, nanohorns, nanoonions, graphene quantum dots, carbon dots or carbon nanoparticles) have ever been used for imaging/tracking of viruses/ viral protein/ viral genome in vivo/ in vitro. I have read articles where they have been used in conjunction with viruses for imaging tumours or for treatment of diseases with drugs but I am not looking for them. I have been scouring for research works in this area but all I could find were metallic nanoparticles, semiconductor quantum dots and very rarly some polymeric/dendritic materials. However, I could find no literature on carbon allotrope based nanomaterials for virus imaging.
I would like to infuse a drug (CNO) and monitor neural activity in the same brain region using calcium imaging (fiber photometry) in mice.
Ideally in the in vivo behaving mouse. But I could start with anesthesized mice.
N.B. I can inject CNO, wait 1/2h, and then run fiber photometry.
I have been using cannulas from Plastics One (Invivo1) for local CNO infusions. Typically in the lab my colleagues have been using Doric fiber photometry systems.
Did anybody ever combine the two ? I see Doric has optogenetic/drug infusion dual systems but I couldnt find fiberphotometry/drug infusion dual systems.
Or maybe I should just make it myself...?
Any input or ideas will be highly appreciated !
I have synthesized some nanoparticles functionalized with antigen and I want to inject them into mice to image their bio-distribution. What are the optimal amount and concentration of the nanoparticle suspension to be injected into the mice? Thanks.
My transgenic mice have more norepinephrine release due to increase in the sympathetic nerve activity which would lead to more vascular smooth muscle contraction. What would be the best technique to measure the vascular reactivity (maybe blood flow in this case) in vivo. I am using both thoracic aorta and mesenteric resistance arteries.
We are experiencing a problem after our cranial window surgeries where after about 2 weeks there is tissue growth that occurs which makes imaging through the windows very difficult with two photon imaging. Also the cranial windows are frequently coming loose or falling off entirely. When we switched from Krazy Glue to Metabond we found that the cranial windows no longer fell off but the same tissue growth still occurred. This seems to indicate that the tissue growth itself was pushing the windows off. Another sign of this tissue growth is an apparent gap develops between the surface of the brain and the window. When we take a close look at the tissue that grows we see it is white in color and seems to grow both over the surrounding skull and over the brain itself (see the following link of the tissue regrowth right after the window has fallen off: http://imgur.com/a/j1cEd). This process seems to be independent of dura removal as the growth happens whether we remove dura or not. We sterilize all tools before use and try to remove as much of the surrounding tissue as possible before installing the cranial window. We're wondering if anyone else has encountered such problems before and if there may be any fixes.
Thanks for any help.
Hi, I am now working on establishing mouse model by Hepa1-6 cell line. 7 days after subcutaneously injection into male C57BL/6J mouse with 5*106 Hepa1-6 cells, solid tumor was established (~0.5cm). However, the tumor was cleared at day 14. I also did intrahepatic injection, no tumor was detected by in vivo image (with luciferase label). Do anyone knows how to establish in vivo model by the Hepa1-6 in C57BL/6J mouse (s.c or i.h)?Is there anything I have to pay special attention? I plan to test my potential anti tumor drug in the model.
Many many thanks.
What do we know about the interaction of cancer cells with murine host cells? And how are murine host cells replaced by cancer cells?
Attached are two coronal MRI images of a mouse lying on its side, with the left image taken 2mm lower than the right one. Can anyone tell me if the part circled by the red line is the hindlimb of the mouse? If not, what is it (some organ?..)? And is the part indicated by the blue square the abdomen? I am new to in vivo MRI images but this must be very obvious to someone with experience. Thanks!
I want to check the Oxidative Burst activity of Neutrophils in a melanoma tumor with in vivo 2 photon microscopy, and I came across the Amplex red reagent, which is said to be a good probe for extracellular H2O2, but I didn't see a reference to it's use in vivo. Does anyone know if it is O.K to use it In vivo?
What is the best method to do it?
Rats will be fed with high calcium diet. I want to measure where the calcium is deposited over time
Radioactive Calcium 45, 19F? I am really lost about the best method!
I'm working on a hypothesis which require in vivo luciferase detection using bioluminescence imaging. But I'm not sure if it can be done in vivo for humans.
I have a Alzheimer disease mouse model and I plan to inject 2 different AAVs as described below:
1. neuron specific promoter-EGFP-WPRE-SpA -AAV2 serotype
2. astrocyte specific promoter-RFP-H1 promoter-target gene shRNA-WPRE-SpA -AAV5 serotype.
I am planning to inject them simultaneously into the cortex of AD mouse model and carry out 2-photon imaging to examine how target gene knockdown in astrocytes influences neuronal number, pathological abnormalities development in a longitudinal study.
Has anyone, injected mixture of AAVs with different serotypes and carried out 2-photon imaging?
I have come across articles where 2-photon imaging of one cell type (either neuron or astrocytes but not together) has been carried out.
I have fluorophore with different excitation/emission parameters, also different serotype, can there be any cross-interaction during 2-photon imaging?
Any particular advice would be helpful.
I am looking forward for any valid and practical protocol for "Exosome labeling that allows them to visualize and tracking their biodistribution in vivo. Do you have any suggestion?
I'm going to use mKate system to label the cancer cell lines, but I noticed there are mKate vectors fused with different proteins available online from Evrogen (http://evrogen.com/products/mKate2/mKate2.shtml). Would there be any difference, in terms of imaging for tumour size, for these? Our lab has a stable cell line with nucleus with RFP, probably mKate, so I’m wondering if that would affect the tumour size measurement, comparing to the cell line has mKate fused in cytoplasm.
Currently I am working on mouse model of non-small cell lung cancer (Adenocarcinoma). For in vivo imaging I am using FMT2500 in vivo imaging system. Lung adenocarcinoma is induced in mice by chemical method (by the use of urethane). I want to visualize the tumor through FMT2500 in vivo imaging system. Please suggest some fluorescent dye to me which is compatible with FMT2500 in vivo imaging system.
We are performing experiments with primary cultures of neonatal murine cardiomiocytes. We were able to succesfully establish the culture and we got the cells beating under the microsocope. How amazing is that!
Now we are trying to measure the effect of different drugs in the cells beating rate and we are not sure how to measure it. The methodology in the papers is not very explicit. We just have to count the beats watching directly the cells under the miscroscope? Are we suppossed to film the cells and then count watching the movie? Is there any specialized software? Any answer will be of help.
Thank you very much!
I'm using 35S-methionine to label in vitro translation products, but due to the short half-life of the label I'm wondering whether I could substitute it with 14C-leucine. Promega and other manufacturers describe that one could use either 35S-methionine, 35S-cysteine or 14C-leucine.
Do you have you experience in using 14C-leucine in in vitro translation?
I'm running SDS- and native PAGE. How long exposure time it requires compared to 35S-methionine? I know, it depends, of course, on the number of methionines/leucines in the protein. But in general, if same amount of methionines and leucines, and suggested amount of radiolabel, does 14C-leucine require very long exposure times? My 35S-methionine exposure to phoshoscreen is good after 24h.
I am going to inject breast cancer (MDA-MB231) cells into female balb/c nude mice. I am injecting at the mammary fat pad. How would we ensure that the tumour is growing within or in the mammary fat pad?
Why NIR fluorescent probes are widely used for in vivo imaging ?
Visible range under 650nm is impossible for in vivo imaging ?
I'm really wonder that...
I want to study the distribution of my nanoparticles in vivo, using a fluorescence tomography imager. I would like to know whether using Alexa Flour 488 dye or Alexa Flour 594 dye is suitable for imaging. As I have read that a Near-Infrared Alexa Flour dye is suitable. However, I would like to use the blue or red spectrum during imaging.
I met a trouble to measure in vivo fluorescence images of S.C injected cy 5.5 tagged siRNA using IVIS imaing. The injected cy 5.5-siRnA was monitoed for its targetting to exnografted tumor whereas a strong fluorescence intensity generated by intestines under the same excitation disturbed the location of released cy5.5-siRNA.
Are there some tips to avoid this noise from intestinal organs please?
During my experiments doing in vivo 2p imaging I get some odd background fluorescence around cells infected with AAV - Syn- GCAMP6. I am using a 10x objective with a low Na, but it seems as if the tissue is glowing, when imaged through a cranial window? Any thoughts what this might be? I was thinking this could be arbors? It does not seem as if the point spread is bad- I get good contrast elsewhere (but will calibrate it all the same in the coming weeks)
I will try to upload an example.
While making two-photon microscopy for a mouse brain through cranial window (in vivo), how much power are you using when you are imaging at the deepest (e.g. 500 um, or even below)? How do you know what is too much? (We have Nikon A1R MP, with Ti:Saphire Chameleon from Coherent)
Is there any recommended standard samples for performing tracking measurements (2D)? Before going to any complicated measurements, we want to run some known samples.
We are going to plan for intravital microscopy,but I am unable to find suitable video how to proceed ,can any one provide
I want to find whether thrombin produces more in some disease condition of lung tissue or blood vessel endothelium，but I cannot find any way to test thrombin formation in organic level，not in the blood
i am working on TEA as a substrate for OCT2 in proximal tubules to determine the OCT2 activity in-vivo.. i am injecting TEA i.v and then take blood samples to determine TEA concentration in serum .. i want a recent simple procedure that can be detected spectrophotometrically
I want to record in vivo (with a video device) the variation of vascular diameter of iris vessels (diameters of 50-125 micrometres) in anaesthetized rats without artefacts (removing respiratory movements with a contention device that I also need). I need an optimized optical magnification at least 20-40x (biomicroscope).
APC/T cells isolated from MOG-immunized mice. During in vitro activation and expansion, I would like to transduce T cells with GFP-encoding virus.
How can I stably express GFP in T cells for in-vivo imaging?
I want to see whether a particular stem cell population can differentiate in vivo (into mice or rat model) into functional neurons or not, if injected in brain.
In vivo imaging or sacrifice of animal and then doing IHC ?
I am confused. Please do give your substantial inputs and any new ideas whatever click into your minds.
I am hoping to get some insight into which spectroscopy techniques would likely be ideal for identifying metal ion protein cofactors in living tissue. Coherent anti-Stokes Raman Spectroscopy, or similar Raman methods, seems to be good, but I am not accustomed with it and do not want to start any research in the wrong direction from the get-go. Sensitivity and discrimination are important qualifications, I should add.
In my experiment, the mice were given subcutaneous injections of GFP stable human tumor cells. Mice were also intravenous injection into tail vein of qDot655 labeled B cells. When I want to use in vivo imaging with Carl zess LSM710, the technician told me that qDot can`t use two-photo.Excuse me, do you know whether qDot655 and GFP staining cells can be seen using two-photon? if not, can you recommend other dye which can be seen together with GFP? thank you very much!
We have been working on an anticancer drug delivery system based on PLGA-b-PEG copolymer nanoparticles, and now are looking for colleagues who are expert in cancer animal models and related in vivo experiments. Our preferential tumor model is MCF7 breast cancer. Please contact me for further detailed information. Thank you.
I have recently investigated into the difference of MITF expression patterns among the human melanoma cancer cell lines composed of SK-MEL-28, MeWo, 501MEL, and A375. These cells predominantly express M isoform of MITF and some of them show the relative expression of A isoform. In vitro, none of them exhibit black color, indicating that they are not able to produce melanin in high amounts. However, due to the analysis of a xenograft model in comparison with in vitro model, MITF expression somehow changes with ectopic melanin synthesis in vivo. What kind of molecular signal effectors might influence on the discrepancy in melanin synthesis potential and MITF expression pattern between in vitro and in vivo?
I am using PE10 tubing for femoral artery catheter, getting the arterial blood pressure signal from a pressure transducer (Harvard apparatus). MABP and Heart rate seems to be in normal ranges (60~90 mmHg and 300~500 bpm, respectively), but pulse pressure is too low, 1~4 mmHg. How can I improve pulse pressure detection?
So according to LiCor, the IRdye secondaries should be soluble in water. I added water to the vial of IRdye (680RD) and now blue chunks of insoluble secondary are floating around. Obviously when you use this for imaging, the image is spotted, random, and not very clear. Any suggestions for getting this IRdye to solubilize?
We would like to immobilize live Daphnia for laser scanning microscopy (confocal microscope). Suggestion of a protocol having minimal impact on the animal physiology would be greatly appreciated.
Would like know if there is any instrument other than ICP-MS (inductively coupled plasma mass spec) that I could use to determine the amount of AuNPs taken up by the liver and other organs during in vivo studies. Thanks
I would like to attach MOLM13-cells for fluorescence microscopy. However, they are cultured in suspension and hardly attach to glass, Poly-L-Lysine-coated glass, or the IbiTreat-slides from IBIDI. Has somebody an idea what to do? If you have information about attaching suspension cells in general and not MOLM13 specifically - thats also fine!
Thanks a lot!
I am looking for any protocol related to in vivo preparations (head-restraint, anesthetized) for 2-photon imaging + electrophysiological recording (extracellular, patch-clamp).
Thank you for your advice
Oxygen metabolism is key to eukaryotic life and important for signaling as well as metabolism. Yet, ROS are linked to cancer and aging as well as inflammation. There is also a possible link between stem cells and levels of ROS. With new in vivo imaging, and other methods ROS levels may be more evident at the individual cell level and in response to environmental stresses.
Is it possible to distinguish between silver ions and silver nano-particles distribution in vivo via an imaging method or a combination? Please let me know, for now I only can find x-ray methods for imaging the particle distribution.
When stimulating electrode immerses in perfusion fluid, there is a 50hz noise, how can I get rid of it?
Which system you recomend if a researcher would like to do non-invasive visualization and tracking of cellular and genetic activity within a living organism such as mouse, in real time. A system with bioluminescence and fluorescence!
I am doing in vivo imaging in the kidney of mice and I want to measure the blood pressure of the mice while I am imaging them.
I want to control the anesthesia depth on one hand but also correlate findings from the imaging with the blood pressure on the other hand.
We have an AD instruments Powerlab and the device to measure the blood pressure invasively via the artery. But I would prefer to measure it non-invasively.
Does anybody have experience with blood pressure measurements in unconscious, isoflurane-anesthetized mice? Does the tail cuff systems (as the NIBP from AD instr.) work there reproducible?
I have planned to go for in vivo studies, i want to kno the name of equipment used for this study. whether Fluorescence microscopy with some required accessories???
Do you think this will match the impressive 0.4deg C found in this study, or are there foreseeable additional hurdles?
I would like to stably knockdown my GOI in the E0771 cell line (mouse breast cancer), which I will then inject into mice to establish a tumour-bearing model. However, I have never done this before and also don't know a lot about the techniques involved. So far, I figured out that I need to transfect my cells with a viral vector expressing either shRNA or miRNA targeting my GOI, and that the viral vector I use needs to be able to integrate into the host genome.
Is this correct? What would be the best option for me to use? What steps/techniques does this procedure require (to generate a vector like capable of this)?
The vector also needs to have some kind of reporter in as well (eg GFP) because I need to be able to visualize tumour growth and metastasis over time (live animal imaging).
I want to measure cytoplasmic ph, in E.coli, under several conditions in-vivo. To this end I would like to use cells expressing a plasmid harboring PH-sensitive GFP variant.
Does anyone have a plasmid like that and is willing to send it to my lab?
alternatively, can anyone please point me to a plasmid repository (other than 'addgene') that might have something like that in stock?
We're performing bioluminescent assays on 5-week old nude mice that have undergone orthotopic xenografts. After anesthesia by isoflurane and retro-orbital injection of D-luciferin, we're seeing unusual mortality events. We're told by imaging staff that deaths are rare during this procedure, but that few researchers use nude mice along with retro-orbital injection of luciferin.
We'd like to minimize loss of mice in these procedures, does anyone have experience with in-vivo luciferase assays that might give us a clue?
I would like to perform an assay for skin dendritic cells tracking towards the draining lymph node and I was wondering if someone have an optimized protocol for FITC or CFSE painting of the skin? Thank you!
If this is possible, what marks or filters are needed for this?
What would you advise for in-vivo two photon calcium imaging between Fura-2 and Oregon Green 488-BAPTA? I cannot do ratiometric analysis because the set-up does not allow it. Any suggestions?
We thought that our university had access to an upright microscope for cranial window experiments in rodents, but as it turns out, we don't have one. Luckily, we will receive a Leica SP8 in a few days but, since it is inverted, we are unsure about how to build a head fixation system that allows for imaging without any unnecessary discomfort for the rodent. If anyone has used or seen a setup like this, I would appreciate some advice and information about the design.
On an upright microscope we would have used the design provided by Hefendehl et al. (Repeatable target localization for long-term in vivo imaging of mice with 2-photon microscopy. J Neurosci Methods, 2012. 205(2): p. 357-63.)
Is there anyone who has tried to label cells with Hoechst or similar in living invertebrates? I am thinking about labeling the nuclei in the brain of a living insect. Has anyone ever tried it?
I am working on a non-model animal and I am searching for in vivo labelling tools. In particular I would like to label cell nuclei (mainly astrocytes and neurons). Is there any specific or cell type-specific dye that I can add ton the brain under the microscope in vivo?
How would you stain glial cells in bees for in vivo imaging? Would SR101 work?