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We narrowed to 468 results for: SEC

Showing: 1 - 20 of 468 results

  1. Even more elegant: Single injection CRISPR/Cas9 in C. elegans

    Type
    Blog Post
    Published
    July 7, 2015, 3:36 p.m.
    ...cassette (SEC). To make it possible fluorescently label proteins in C. elegans, they inserted the SEC into...their Roller phenotype. If SEC is inserted at the 5’ end of a gene, the SEC separates the targeted gene...to remove the SEC and after 5-6 days of growth, wild-type animals that have undergone SEC excision are ...the Genetics Society of America. To use this SEC-based system, first design and clone 500-700 bp homology...homology arms for your gene of interest into the FP-SEC vector using Gibson assembly. This construct will... attempt for 6/7 genes. The data are clear - the SEC-based system is both simple to use and highly efficient...genome modifications, including targeted mutations. SEC-based systems may prove effective in other species...

  2. Antibodies 101: Secondary Antibodies

    Type
    Blog Post
    Published
    Dec. 21, 2021, 2:15 p.m.
    ...about secondary antibodies. While a primary antibody binds to the protein of interest, a secondary antibody...conjugated to the secondary antibody instead of the primary antibody. Since multiple secondary antibodies can...with BioRender.com.    How do secondary antibodies work?  Secondary antibodies are antibodies generated...While most secondary antibodies recognize broad isotype classes, it is possible to get secondary antibodies... conjugated secondary antibody, which will bind to the primary antibody. Perform a second series of wash.... Primary antibodies are represented in blue; secondary antibodies are represented in purple; signaling...antibody’s Y) is large enough to allow multiple secondary antibodies to bind to a primary antibody without...

  4. Plasmids 101: Secondary Nanobody Toolbox

    Type
    Blog Post
    Published
    Feb. 27, 2018, 2:04 p.m.
    ...techniques have in common? They all typically require secondary antibodies, frequently of the mouse or rabbit ...Görlich lab and their anti-mouse and -rabbit IgG secondary nanobodies toolbox. Nanobodies are like tiny antibodies...the following to occur: A) Multiple polyclonal secondary IgGs can bind to different epitopes of a primary...antibodies are large and bivalent, primary and secondary antibodies cannot be incubated together without... least from different IgG subclasses, so that secondary antibodies exclusively recognize only one primary...yielding similar signal strength to a standard IgG secondary antibody (Pleiner et al figures 4A-C). And since...targets (Pleiner et al figure 4D). Unlike large secondary antibodies, small nanobodies do not oligomerize...

  6. Production of Virus in Insect Versus Mammalian Cells

    Type
    Blog Post
    Published
    Nov. 5, 2024, 8:32 p.m.
    ...purification using qPCR or digital droplet PCR. Insect cells Insect cells are a less common option for the generation... viruses that specifically infect insects. This is what makes insect cells a popular choice for protein...Mammalian versus insect cells for viral production Consideration Mammalian cells Insect cells Viruses...different cell types, including plants, mammals, and insects. In this post, we will compare two of the most ...most widely used systems, mammalian and insect cells. Whether you’re new to in-house virus production or ...passage (transfection). This differs from typical insect cell preparations, which we talk about below. Processing...protein production. The most commonly used cells in insect viral production are Sf9 cells, which are derived...

  7. Avoiding the Dark Side of Fluorescent Protein Fusions with mOX FPs

    Type
    Blog Post
    Published
    Oct. 27, 2015, 3 p.m.
    ... in oxidizing environments when targeted via the Sec translocon." Traffic 12.5 (2011): 543-548. Pubmed...compose the secretory pathway, endocytic vesicles, mitochondria, lysosomes or they are secreted into the ... blue fluorescent protein, secBFP2, for studies in the eukaryotic secretory pathway."Biochemical and biophysical...residues form inappropriate disulfide bonds in the secretory pathway (Figure 1) [4,6–8]. Additionally, FPs ...and a sequence intended to localize them to the secretory pathway, a dark pool that is mislocalized to the...misfolded, blocked from proceeding through the secretory pathway, and retained in the ER. FPs have been...variety of cellular compartments including the secretory pathway (Figure 2), the inner membrane space of...

  9. Antibodies 101: Choosing the Right Isotype

    Type
    Blog Post
    Published
    April 6, 2023, 3 p.m.
    ...use a highly specific secondary antibody, rather than a broadly  reactive secondary (Fig. 1). For example... a broadly reactive secondary antibody to increase the chance of primary/secondary antibody binding and... a distinctly conjugated secondary antibody. Consider too that some secondary antibodies can cross react... IgG2a cross-adsorbed secondary and a Goat anti-rat IgG1cross-adsorbed secondary would limit species cross-reactivity...Benefits and Pitfalls of Secondary Antibodies: Why Choosing the Right Secondary Is of Primary Importance...applications. The desired specificity of your secondary antibody depends on whether you are using poly...poly- or monoclonal primary antibodies. Some secondary antibodies are broad and recognize multiple isotypes...

  10. Antibodies 101: Avoiding the Mouse on Mouse Mess in IHC

    Type
    Blog Post
    Published
    Sept. 21, 2023, 1:15 p.m.
    ...by mixing your primary and secondary antibodies, blocking excess secondary with serum from the same species...apply your primary and secondary antibodies as normal. Ideally, your secondary will only bind to your ...the same host species as your secondary antibody since 1) your secondary should not recognize Fab fragments...your primary and secondary antibodies before adding them to your sample so the secondary antibodies aren...antibodies - one against your target of interest and a second against the first antibody. When endogenous IgGs...species as your primary antibody are present, your secondary antibody recognizes both the endogenous IgGs and...staining with no problem! But to be sure that your secondary is not binding endogenous IgGs, you should include...

  11. A Control for All Seasons

    Type
    Blog Post
    Published
    Oct. 5, 2023, 1:15 p.m.
    ...each primary is omitted but all secondaries included to uncover secondary antibody cross-reactivity. ...antigen-binding site.      Secondary antibody specificity  In indirect staining approaches, secondary antibody specificity...and the use of isotype-specific secondary antibodies.  Baseline secondary signaling levels In addition to... substituting for a more specific secondary antibody. The secondary antibody control should be included...staining with multiple primary and secondary antibodies. Ideally, secondary antibodies will be highly specific...signal from your anti-Mouse IgG2a secondary antibody, then that secondary antibody is reacting non-specifically...experimental group, with the new kinase inhibitor and a second group, the positive control group, with an alternative...

  12. Antibodies 101: Multiplex Immunofluorescence

    Type
    Blog Post
    Published
    May 23, 2023, 1:15 p.m.
    ... is what conjugated secondary antibodies you have available. If the only secondary antibodies you have... in your sample, then a fluorescently labeled secondary antibody binds that first antibody, then you use...approaches is to use sequential cycles of primary and secondary antibody labeling and imaging, repeating for each...mixture of primary antibodies and appropriate secondary antibodies. The general process is similar to ...imaged at a time, inactivating the fluorescent secondary antibodies before labeling the next target. B)...antibodies are combined in one step and all the secondary antibodies are combined in one step. As with...frozen or formalin-fixed, paraffin-embedded tissue sections, methanol- or paraformaldehyde-fixed cell samples...

  13. RANbodies: Reporter Nanobody Fusions

    Type
    Blog Post
    Published
    April 10, 2018, 12:56 p.m.
    ...primary/labeled secondary Less sensitive than HRP-RANbody staining Requires secondary antibody staining...primary/labeled secondary   Less sensitive than HRP-RANbody staining Requires secondary antibody staining... RANbodies are expressed in 293T cells and are secreted from cells due to their IgGκ signal peptide sequence...three reporters stain cultured cells and tissue sections, some reporters are better suited for particular...summary. HRP HRP tagged RANbodies don’t require a secondary antibody for detection. Instead they are detected... signal over staining with a primary/ labeled secondary antibody and a ≥100-fold increase in signal over...directly labeled primary or a primary/labeled secondary antibody are used to detect additional antigens...

  14. Antibodies 101: The Four ELISAs and When to Use Them

    Type
    Blog Post
    Published
    July 25, 2024, 1:15 p.m.
    ...no need for a secondary antibody incubation step, saving time. Moreover, since secondary antibodies can...primary antibody and a conjugated secondary antibody. Incorporating a secondary antibody increases the sensitivity...antigen and then a reporter enzyme conjugated secondary antibody binds to the primary antibody. In a sandwich...that binds to the antigen in a sample and then a second detection antibody binds. The detection antibody...conjugated with a reporter enzyme or a conjugated secondary antibody can be used. In a competitive ELISA a...also lack the signal amplification afforded by secondary antibodies and thus are not suitable for low abundance...sensitivity of the assay, since multiple secondary antibodies will bind to a single primary antibody and ...

  15. Antibodies 101: Introduction to Immunofluorescence

    Type
    Blog Post
    Published
    Nov. 16, 2021, 2:15 p.m.
    ...common is that secondary antibodies are generally polyclonal, which means that multiple secondary molecules...and incubation time. Secondary Antibody Unsurprisingly, this is the second antibody you apply, which...indirect IF requires two antibodies: a primary and secondary. For indirect IF, the primary antibody targets...attached to a fluorescent molecule, instead a secondary antibody that targets the primary is conjugated...it is more efficient to generate a handful of secondary antibodies that target common IgGs and then conjugate.... It is common for labs to keep a few trusted secondary antibodies on hand to mix and match with a larger...Speaking of different colors, having a palette of secondary antibodies in a few different species and colors...

  16. Antibodies 101: Chimeric Antibodies

    Type
    Blog Post
    Published
    April 25, 2023, 1:15 p.m.
    ...anti-rat IgG1 secondary to detect Protein A and green fluorophore-tagged anti-mouse IgG2a secondary to detect...IgG1 secondary antibody to bind to the anti-Protein A antibody and an anti-mouse IgG2a secondary antibody...constraining using the indirect method since a secondary antibody reactive to a certain isotype will bind...antibodies that are the same isotype then the secondary antibody will bind to both and you will not be...the same isotype and will react with the same secondary. If you were to change one of the antibody’s isotypes...isotypes, however, you could then use 2 different secondaries for your antibody pair and visualize both targets...multiplexed indirect staining methods since the secondary antibody used will react to both primary antibodies...

  17. Which Fluorescence Microscopy Techniques is Best for Me?

    Type
    Blog Post
    Published
    Oct. 10, 2017, 1:57 p.m.
    ...microscopy technique to incorporate optical sectioning. Optical sectioning refers to the ability to extract light...) tissue sections and 3D cultures These samples do not necessarily require optical sectioning, but open...this, the use of multiple fluorophores can add a second layer of contrast, color contrast, to an image ...volumes to enhance spatial contrast (i.e. optical sectioning (2) or super-resolution (3). Fluorescence microscopy...establish for this to happen.Using red light has a second advantage: red light penetrates deeper into biological... on the sample, a temporal resolution on the millisecond scale may be needed. Most commonly these experiments... of the coverslip (4). Essentially, an optical section is achieved that boosts contrast by filtering out...

  18. Plasmids 101: Protein Expression

    Type
    Blog Post
    Published
    June 7, 2018, 1:17 p.m.
    ...conditions compared to other systems. Insect expression systems Insect cells can also be used to produce ...are two types of insect expression systems; baculovirus infected and non-lytic insect cells. Baculovirus...genes integrated into the insect cell genome. Both of these types of insect expression systems can be ...stable proteins. Protein expression refers to the second step of this process: the synthesis of proteins...production and purification. These include mammalian, insect, bacterial, plant, yeast and cell free expression...halts protein production, but there are non-lytic insect cell expression systems (sf9, Sf21, Hi-5 cells)...of large amounts of protein. Some drawbacks of insect cell expression systems include that virus production...

  19. A Guide to Designing a Scientific Poster: Content, Layout, and More

    Type
    Blog Post
    Published
    Oct. 22, 2019, 12:20 p.m.
    ...acknowledgements sections. Some other additions you might wish to include are a references section, contact information...styles; one for your section titles, and a different one for the body of those sections. This strategy helps...poster. While a poster generally contains the same sections as a primary research article, it’s important ...common poster size, but this can vary as well.  Sections of your poster In almost all scientific posters...creative use of a footer in your poster. This is a section where you can insert your references and contact...font here to be much smaller than in other poster sections, so you can include a lot more. Font size: Making...helps your sections stand out, and makes the entire poster more navigable.  Choose your content wisely...

  20. Antibodies 101: Selecting the Right Antibody

    Type
    Blog Post
    Published
    March 15, 2022, 1:15 p.m.
    ...in a similar way as you plan to. To secondary or not to secondary?  The application will also determine...primary-conjugated) or indirect (secondary) methods. In those cases, remember that a secondary should be used when ...substrate, such as a fluorophore, or a conjugated secondary antibody that will bind to your primary antibody... the precise location of a protein in a tissue section then a directly-conjugated antibody will provide...immune complexes are smaller than a primary:secondaries:conjugates complex. If you are trying to visualize...protein in low abundance on a Western blot, a secondary antibody is ideal as it will help to amplify the...you can choose different fluorophore-conjugated secondary antibodies specific to each isotype or species...
Showing: 1 - 20 of 468 results