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We narrowed to 475 results for: FLU

Showing: 61 - 80 of 475 results

  1. Immunocytochemistry

    Type
    Protocol
    ...primary antibody against a target protein and a fluorescent secondary antibody....primary antibody against a target protein and a fluorescent secondary antibody. This protocol outlines the... tips and pipettes Rocking platform Tweezers Fluorescent microscope 0.5–10 µL single channel pipette 2...500 µL PBS on a rocking platform. Dilute the fluorescently-labeled secondary antibody to the desired concentration...generally ranges from 1-10 µg/mL. Add 500 µL fluorescently-labeled secondary antibody to the wells and ...cell labeling on a microscope with appropriate fluorescent filters. Tips and Troubleshooting The optimal...

  2. AAV Production in HEK293 Cells

    Type
    Protocol
    ...maintenance phase and do not allow cells to reach 100% confluence (80–90% is ideal). Pass and plate the cells the...from 2 x T-175 flasks. Cells should be at ~80% confluence. For each T-175 flask: Aspirate culture media...resuspend cells from the CS2. Cells should be at ~80% confluence. Aspirate culture media and rinse once with 60...Sample Data Figure 1: HEK293T cells at various confluencies. This AAV production protocol should be started...started with cells that are ~80% confluent (center panel). These cells were imaged with a 10X objective ...

  3. General Transfection

    Type
    Protocol
    ...with fresh media Day 3 or more (am): Observe fluorescence, harvest cells, or perform your experiment Equipment...batch of PEI is prepared, transfect cells with a fluorescent plasmid using a variety of ratios. Check the ...efficiencies as can be seen here by the amount of green fluorescent protein expression (green in the right panels...

  4. Virus Protocol - Generating Stable Cell Lines

    Type
    Protocol
    ...the individual wells become confluent, expand into larger vessels. A confluent well of a 6-well dish can...can be expanded into a 10 cm dish. A confluent 10 cm dish can be expanded into two 75 cm 2 flasks, etc....

  5. Isolating a Monoclonal Cell Population by Limiting Dilution

    Type
    Protocol
    ...the next day they will be approximately 50–60% confluent. For 293T cells this is about 2 × 10 6 cells in...Do not use the medium if the cells are overly confluent or the medium is acidified (i.e., the medium appears...cells have expanded but before they become over-confluent, trypsinize the cells and expand them to larger...

  6. Protocol - pLKO.1 – TRC Cloning Vector

    Type
    Protocol
    ...cells from incubator. The cells should be 50-80% confluent and in DMEM that does not contain antibiotics....The target cells should be approximately 80-90% confluent. c. Dilute puromycin in the preferred culture ...: b. Target cells should be approximately 70% confluent. Change to fresh culture media containing 8 μg...

  9. Lentivirus Production

    Type
    Protocol
    ...batch of PEI is prepared, transfect cells with a fluorescent plasmid using a variety of ratios. Check the ...

  10. CRISPR Library Amplification

    Type
    Protocol
    ...different concentrations of DNA. In our hands fluorescent dye methods tend to report lower values than ...

  11. Lentivirus ddPCR Titration

    Type
    Protocol
    ... Colony Formation Titration Assay Lentivirus Fluorescence Titration Assay AAV Titration by qPCR Addgene...

  12. Antibodies 101: Designing Your First Flow Panel

    Type
    Blog Post
    ...  APCAlexa Fluor 647   Alexa Fluor 700APC-H7APC-Cy7   Many factors can influence the level of... you'll need to pick a fluorophore for each target and ensure the fluorophores, together, are able to ...reporter's fluorescence. Another way to label proteins is through treating cells with fluorophore-conjugated...dead cells. The combination of antibody-fluorophores, fluorescent reporters, and/or dye stains used in your...outlining a few commonly used fluorophores and their emission spectra. Some fluorophores carry their emission ...and RB545. Fluorophores can also come in combination of two conjugated single fluorophores, like APC-Cy7...respective colors of a few commonly used fluorophores. Each fluorophore has a defined emission maximum. Note...

  13. Technologies Enabled by NanoLuc® Luciferase

    Type
    Blog Post
    ...fusion protein called CalFlux VNT. CalFlux VNT is composed of Venus fluorescent protein, the Troponin ...NLuc to fluorescent proteins to make better reporters and the second couples NLuc to fluorescent proteins...better biosensors. NanoLuc®-fluorescent protein fusions for imaging Fluorescent proteins are fantastic imaging...couple Nluc with a red-shifted fluor. Here, NLuc internally excites the fluor through a process known as ...BRET). The fluor then emits light at a tissue-penetrating wavelength above 600 nm. LumiFluors Schaub, F.X...more penetrative orange fluorescent protein fusion (OgNLuc). Both fluorescent proteins can be excited ... the lab has dubbed these fusions LumiFluor Reporters. The LumiFluor reporter constructs were designed...

  14. Antibodies 101: Flow Cytometry

    Type
    Blog Post
    ...the associated fluorophore or stained with the associated fluorochrome, emits a fluorescent signal that ...the level of fluorescence. A cell expressing high levels of the fluorophore or fluorochrome-stained protein...antibody conjugated fluorophores Fluorescent antibodies get around the challenges of fluorescent protein fusions...such as fluorescent microscopy must be used. To learn more about the most suitable fluorescent microscopy... have a variety of fluorescent light sources. When interrogated with a fluorescent light source, a cell... to a fluorochrome. In indirect staining a primary antibody binds to a target and a fluorochrome-conjugated... panel of “fluorescence-minus one” controls, samples stained with all of the fluorochromes in the experiment...

  15. Tips for Using FRET in Your Experiments

    Type
    Blog Post
    ...non-radiatively from an excited molecular fluorophore (the donor) to another fluorophore (the acceptor) by means of ...the two fluorophores used are in close vicinity. Thus, the distance between the two fluorophores and their... of a fluorophore pair, donor:acceptor stoichiometry, and cross-talk between the two fluorophore colours... filters chosen for the donor fluorescence and the acceptor fluorescence. Additionally, this method could... the fluorescence decay time of the donor. When FRET occurs between the pairs, donor fluorescence is quenched...factors, such as pH or autofluorescence background, can change the fluorescence decay time and have to...course talking about the now well-known FRET, aka Fluorescence (Förster) Resonance Energy Transfer, technique...

  16. Antibodies 101: Conventional vs Spectral Flow Cytometry

    Type
    Blog Post
    ...for autofluorescence extraction Cells produce a low level of fluorescence called autofluorescence (you...to identify a fluorophore, spectral flow cytometers use all detectors for all fluorophores. This allows...tool called “autofluorescence extraction” which allows the user to use the autofluorescence in a sample...sample as a fluorescent marker. Given that the level of autofluorescence across various tissue types varies...you wish to use the autofluorescence extraction feature. Although autofluorescence extraction is an optional...samples stained with a single fluorophore. They’re required for every fluorophore you use in your panel, whether... the spectral signature of a fluorophore itself from the autofluorescence of the beads or cells. Overall...

  17. Hot Biosensors 2022: Year-End Roundup

    Type
    Blog Post
    ... blog Fluorescent Proteins 101: Fluorescent Biosensors Resources on Addgene.org Fluorescent protein...area with lots of bright new plasmids is our Fluorescent Biosensors Collection — scientists are always...measure! Let’s take a look at some of the hottest fluorescent biosensors new to Addgene this year. Feel the...Hiroshi Hibino and colleagues devised a “Modular fluorescence unit fused with TRansmembrane region-to-IntrAcellular...region-to-IntrAcellular loop linkers” (MTRIA for short), where a fluorescent protein is inserted into one of the loop regions...  Find MTRIA sensors here!   Ino D, et al. A fluorescent sensor for real-time measurement of extracellular...co-expressing a cytosolic ZtSH2 reporter, they observe fluorescence localizes to the plasma membrane only when the...

  18. CRISPRainbow and Genome Visualization

    Type
    Blog Post
    ...More about Fluorescent Protein-Based Tools Learn Some of the Pitfalls When Using Fluorescent Proteins Control...Resources on Asddgene.org Find Fluorescent Protein Plasmids Find Fluorescent Protein Kits Find Empty Backbones...historically been difficult, as techniques like fluorescent in situ hybridization (FISH) and chromosome capture...be used in vivo. Some researchers have used fluorescently tagged DNA-binding proteins to label certain...chromosome. This image represents the composite of fluorescent measurements from red, blue, and green channels...targeting capacity. By using a gRNA to target fluorescently labeled dCas9 to a given genomic locus, you ... thereby decreasing the amount of background fluorescence from unbound GFP-dCas9. With these improvements...

  19. Live and Let Dye: Self-Labeling Protein Tags

    Type
    Blog Post
    ...years — a green fluorescent protein, a red fluorescent protein, a fancy new green fluorescent protein, some...new-and-improved fluorescent protein you want to try, just swap in a new fluorescent ligand and see the...purification tags, another version with that same red fluorescent protein somebody made because they forgot it ...of chemical dyes provide many advantages over fluorescent proteins, opening up new applications.   ...strategies. The green structure represents a fluorophore or other useful ligand, gray structure is the...biarsenical dyes such as FlAsH (green, based on fluorescein) or ReAsH (red, based on resorufin). SNAP-tag...dihydrofolate reductase (eDHFR) could be labeled with fluorescent derivatives of its substrate trimpethoprim (TMP-tag...

  20. In Living Color: The Skinny on In Vivo Imaging Tools

    Type
    Blog Post
    ...than any fluorescent protein earlier described. “Until our proteins, there were lots of fluorescent proteins... If you start poking around on Addgene’s Fluorescent Protein Guide to In Vivo Imaging, you’ll pretty ...good reason. We all know scientists have used fluorescent proteins to observe what’s happening inside cells... of decades. Green is the classic color, but fluorescent proteins are available in a variety of hues. ...unimpeded. The challenge then was to develop fluorescent proteins that would fall within that near-infrared... from jellyfishes and corals, but all of them fluoresce outside the transparency window of mammalian tissues... Verkhusha said. “We developed near-infrared fluorescent proteins within this transparency window, so ...
Showing: 61 - 80 of 475 results