...you turn up the brightness in your fluorescent imaging experiments. Fluorescent protein fusions One of...expressed at low levels, you have to increase your imaging time to get enough signal. This workaround risks...phototoxicity and eliminates the possibility of long-term imaging studies. If you instead overexpress your protein...examined the power of SunTag for single molecule imaging, finding that plasma membrane-targeted CAAX-SunTag...power of SunTag, they attempted single-molecule imaging deep inside the cell, in the nucleus and cytoplasm...mitoNEET-SunTag-GFP. As expected, they obtained bright images of mitochondria without organelle toxicity. First...protein aggregation, it’s recommended for most imaging applications. What can you do with SunTag? Tanenbaum...

...for in vivo and in vitro imaging. Saito et al. used this Nano-lantern to image labeled tumors in freely...associated with luminescent imaging: A luminescent substrate is required for imaging, and it is important ...Fluorescent imaging techniques have become indispensable tools for molecular and cell biologists over...intensity of luminescent proteins. To make luminescent imaging a reality, Addgene depositor Takeharu Nagai and...scientists from using chemiluminescent proteins for imaging, but it has precluded widespread adoption of the...coelenterazine, with increased sensitivity and faster imaging compared to previous publications. They also developed...photobleaching/phototoxicity, they successfully imaged multiple subcellular compartments over several ...

...response to a ~25 mT magnet. Gray regions indicate times when the magnetic field is on. Image reproduced from...response to a ~10 mT magnet. Gray regions indicate times when the magnetic field is on. Image reproduced from...of E. coli expressing MagLOV while a magnet is waved under the sample. The imaged area measures approximately...whenever the magnet approached the sample (by less than 1%) and recovered after the magnet was removed ...75% in response to the magnetic field (Figure 2); they named this variant MagLOV. Andrew was thrilled to... to see that MagLOV’s “astonishing” magnetoresponse was clearly visible whether expressed in E. coli, ...variants of MagLOV will be possible with other engineering approaches and that magnetosensitive tools can...

...Addgene’s Fluorescent Protein Guide to In Vivo Imaging, you’ll pretty quickly notice the name Vladislav... light in the near-infrared portion of the electromagnetic spectrum – the spectral region in which mammalian...FPs), called PAiRFP1 and PAiRFP2, to the in vivo imaging toolbox, which increase their fluorescence upon...as photoacoustic contrast agents for two-color imaging in animals. The approach relies on ultrasound waves...as opposed to light to produce higher resolution images in vivo. “One of the reasons we can’t localize ... a result, ultrasound enables higher resolution images at depths up to eight millimeters – not single ... near-infrared fluorescent protein for in vivo imaging. Filonov et al (Nat Biotechnol. 2011 Jul 17. doi...

...Thorn of the Nikon Imaging Center at UCSF. A common requirement for live cell imaging experiments is the...emission spectra differ sufficiently for them to be imaged in distinct fluorescent channels on the microscope...many fluorescent protein combinations that can be imaged together, but this also means that the choice of...choosing fluorescent proteins for your multi-color imaging experiment is to be aware of what fluorescent proteins...proteins To choose a set of fluorescent proteins to be imaged together, you will need to consider the same factors... distinguished from one another and that can be imaged with the optics on the microscope(s) you intend... 474 nm) and YFP (ex 514 nm / em 527 nm) can be imaged together but CFP and GFP (ex 488 nm / em 507 nm...

... anyone performing multiplexed imaging, not just IBEX. The IBEX Imaging Community’s vision is an open,...IBEX is a highly multiplexed tissue imaging method that uses iterative staining and chemical bleaching...bleaching to allow for high-resolution imaging without degrading the tissue (Radtke et al., 2020, 2022). I do... do mean highly multiplexed: you can image over 65 different parameters using this method (see Movie S8... experiments!)     Figure 1: The IBEX Imaging Community front page. That’s why scientists... including 61 fluorescent probes, representing imaging from 47 different tissues. Researchers upload and...experiments. Researchers can then visit the IBEX imaging community discussion board to dive into conversations...

...Deeper: Biomolecular Tools for Acoustic and Magnetic Imaging and Control of Cellular Function. Biochemistry...other hand, is a widely available and inexpensive imaging modality with good tissue penetration depth as ...enable cells to be visualized under ultrasound imaging. Expression of ARGs results in the production of...background tissue unless destructive ultrasound imaging was used. We set out to develop next-generation...L-arabinose intraperitoneally and wait 24 hours before imaging with ultrasound to observe gas vesicle-specific...TEM, and purified GVs under TEM. (c) Ultrasound imaging of tumor colonization by bARGSer-expressing E. ... tumors was overlaid with the anatomical B-mode images in greyscale.   ARG expression in mammalian...

...in ultrasound imaging. Ultrasound is an attractive imaging platform for in vivo imaging experiments because...and widely available imaging platform: ultrasound. Challenges of in vivo imaging with existing bacterial...Limited Depth of Signal/Imaging: Fluorescent reporters are not as well suited for imaging organs located deep...ARG’s: in vivo imaging. Specifically, arg1 expressing E. coli aided in ultrasound imaging of the mouse ... injected into the mouse colon, imaged via ultrasound, and the images compared to mice injected with ECN...anatomical context/spatial localization: In vivo imaging with fluorescent or bioluminescent reporters usually...and then arg1 vesicles. By comparing ultrasound images pre-collapse and post-collapse of arg2 and then...

...newly formed proteins.  Limitations with GFP in imaging  While GFP has been an immensely powerful tool ...protein maturation times which limits its use for imaging a protein’s entire life-cycle. This makes it nearly...nearly impossible for scientists to use GFP to image nascent proteins. In addition GFP tags are large,...attached to a protein, making them hard to amplify and image for long periods of time.  Are antibody based probes...used to amplify fluorescent signals in in vivo imaging: the 10xHA tag increased the fluorescent signal... mCherry, HaloTag and SNAP-tag for multi-color imaging. Find the HA frankenbody tool-kit at Addgene to...proteins.  Lastly, the HA frankenbody can be used for imaging in whole organisms. The researchers used the HA...

...and their transparency allows for whole organism imaging. They created an endogenous ApoB-NanoLuc reporter... transmit electrical information. Such voltage imaging can be performed in intact animals and offers many...

... at Addgene. Sherf, et al. 1994. Promega Notes Mag. Giannakis, et al. BioTechniques. 2003. PubMed ...Nano-lanterns for multicolor luminescence imaging  Fluorescent imaging techniques have become indispensable...):198-200. SunTag system for single molecule imaging and inducible gene expression Brilliant technologies...have adopted and employed this concept in both imaging and inducible gene expression studies, which are...advanced technology in the fields of single molecule imaging and inducible gene expression by creating the SunTag...system, cleverly termed TRICK, for Translation RNA Imaging by Coat protein Knock-off, is made up of three ...developed a CRISPR-based multicolor tool enabling the imaging of multiple endogenous genomic loci simultaneously...

...x-ray, or a biologist imaging the smallest structures of life; the key to a great image is contrast. The human...Epi-fluorescence microscopes. (See left image above for an example image). Point Scanning Confocal Microscopy...the speed at which a confocal image can be acquired. Assembling an image pixel by pixel is slow; therefore... challenging samples to image as they require both optical sectioning, imaging over large distances and...can push imaging depths closer to 1 mm. Traditionally, 2-photon has been a very slow imaging technique...Lightsheet designs that allow for multi-view imaging can image thick scattering samples in their entirety...photobleaching when acquiring multiple images over a large axial range. 2-photon imaging, which restricts excitation...

...open the offending red-green image, go to Image>Color>Replace Red with Magenta. Or you can use Color>Split...wash again, then image. But here’s where the “sequential” part comes in. After you image the first target....1 (magenta), anti-AnkyrinG (green), and Hoechst nuclear stain (cyan). Panels C1-C3 show magnified detail...ve captured your images, various software programs will allow you to align the images of each labeled ...maybe you just want to make some really beautiful images? Multiplex IF may be just what you need! How does...of primary and secondary antibody labeling and imaging, repeating for each target. Alternatively, you ...Sequential Multiplex IF, where one target is labeled and imaged at a time, inactivating the fluorescent secondary...

...called Magnetic Isolation and Concentration (MagIC)-cryo-EM that combines enrichment and imaging of proteins...proteins captured on magnetic beads (Arimura et al., 2024). In MagIC-cryo-EM, target proteins (expressed...plasmids for MagIC-Cryo-EM here!   Arimura, Y., Konishi, H.A., Funabiki, H. (2024) MagIC-Cryo-EM: Structural...articles. Here's what you'll find in this post: MagIC-Cryo-EM for enrichment and structure determination...Recombinant antibodies for marking astrocytes MagIC-Cryo-EM for sample enrichment and structure determination...proof-of-concept study) are captured on streptavidin-coated magnetic beads through an assembly of biotinylated SpyTag-SpyCatcher...and minimizes sample loss while also supporting imaging, making it easy to identify dozens or hundreds ...

...resolution imaging, where the power of the technique relies heavily on the same fluorophore being imaged multiple...order to reconstruct an image. The best PA-FPs used for super-resolution imaging usually have the following...Fluorescent Protein Imaging Choosing Your Fluorescent Protein for Multi-Color Imaging Catch Up on Your GFP...would otherwise be missed with conventional FP imaging. PA-FPs are divided into 2 main categories - those...photoactivation - and have enabled sophisticated imaging techniques.     Irreversible photoactivation Irreversible...phototoxic to cells, most scientists working on in vivo imaging models perfer the PA-RFPs over PA-GFP. Another ...fluorescent state make them useful for 2-color imaging with green PA-FPs. Table 2: Properties of Selected...

...Individual images as well as collections of images are easy to download in a .png file format. Images are under...indicate if changes were made to the images. This requirement means the images are probably better suited for...than for journal publications. Google Image Search Google image search is a great tool for searching the...can search for images “Labeled for reuse with modification.” Occasionally Google Image Search results ...results includes images that can’t be reused without attribution, despite filtering for images labeled for reuse...your public library. If you want to use pre-drawn images in your figures Biorender Biorender is like clip...create it. Additionally, you can upload your own images to the Biorender workspace. Free Biorender accounts...

...named it Magneto 2.0. Using Magneto2.0-p2A-mCherry constructs, the lab verified that Magneto2.0 was magnetically...revealed that Magneto 2.0 allows for magnetic remote control of neuronal activation. Find Magneto 2.0 plasmids...is multiplex automated genome engineering (MAGE) (1). MAGE uses recombineering (2) to simultaneously incorporate...use the MAGE method in unmodified bacterial species (4). This set of plasmids (dubbed pORTMAGE) expresses.... Methods 2016. PubMed PMID: 27798611 Magneto 2.0 - A magnetic remote control for the nervous system ... 74308 pcDNA3.0-Magneto2.0-p2A-mCherry Mammalian Expression 74334 pCR8-Magneto2.0-p2A-mCherry Gateway...Gateway 74333 pCR8-Magneto2.0 Gateway 74307 pAAV-CMV-DIO-Magneto2.0-sNRPpA AAV 74306 pAAV-CMV-DIO-TRPV4...

...excitation microscopy is a fluorescence imaging technique that allows imaging of living tissue up to about one... technique and the in vivo live-cell imaging (intravital imaging) studies it’s enabled are revealing the...immune cell mobility than the static microscopy images of chemically coloured lymphoid organs did in the...Two-photon microscopy was the next revolution in cell imaging, giving researchers the ability to monitor cellular...than any other cell type in the body! Intravital imaging powered by two-photon microscopy has uncovered ...infected tissues. Before the use of two-photon imaging, our understanding of pathogen-immune cell interaction...Neutrophils travel to the sites of infection or damage (Peters et al. 2008, Chtanova et al. 2008, Kamenyeva...

...fluorescent protein fusions for imaging Fluorescent proteins are fantastic imaging tools but their use is limited...for imaging. These authors set out to design a better fluorescent protein that could be co-imaged with...of researchers wanting to perform whole-animal imaging, however NLuc’s emission peak is at 460 nm thus...thus making it non-optimal for in vivo imaging. The solution is to couple Nluc with a red-shifted fluor....LumiFluors Schaub, F.X., et al. (3) designed two imaging probes. One consists of NLuc fused to enhanced ... The isolated cells were injected into mice and imaged after i.p. or i.v. administration of the furimazine...of why a BRET reporter can improve deep tissue imaging. The constructs for GpNLuc and OgNLuc are available...