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Cat. No.: AFMB-0.5 (for 0.5ml)

Cat. No.: AFMB-2 (for 2ml)

Description

Anti-Flag Magnetic Beads, also known as Anti-Flag immunomagnetic beads or Flag antibody magnetic beads, are formed by covalently coupling high-quality Flag mouse monoclonal antibodies with nano-sized amino magnetic beads. These beads can specifically bind to proteins containing Flag tags in lysates, serum, ascites, and other samples from animals, plants, or microorganisms. They are used for immunoprecipitation (IP), co-immunoprecipitation (Co-IP), or purification of fusion proteins or protein complexes containing Flag tags.

Flag tag, Myc tag, HA tag, His tag, GST tag, and others are commonly used tags on expression vectors. Fusion expression with these tags allows for convenient detection of the target protein and its interacting proteins, as well as purification of the target protein.

The Flag tag is a peptide consisting of eight amino acid residues (DYKDDDDK). The common forms of Flag tag are Flag and 3X Flag. By recombinant DNA technology, the nucleotide sequence of the Flag tag is linked to the 5' or 3' end of the target gene, resulting in the expression of the target protein with the Flag tag. The Flag tag has the following advantages: it usually does not interact with the target protein and does not significantly affect its function; as a tag protein, the expression, localization, and function of the target gene can be detected using Flag antibody (AF519), Anti-Flag magnetic beads, or Anti-Flag affinity gels (P2271), and the target protein can be purified, immunoprecipitated, or co-immunoprecipitated; the Flag tag fused at the N-terminus can be cleaved by enterokinase (EK) at the DDDK site, resulting in the target protein without the tag. Based on these advantages, the Flag tag has been widely used in various aspects of protein expression, purification, identification, interaction, and function research.

Anti-Flag Magnetic Beads (also known as Anti-DYKDDDDK Magnetic Beads) can specifically bind to fusion proteins with Flag tags. They can be conveniently applied for immunoprecipitation or purification of fusion proteins or protein complexes containing Flag tags using magnetic separation devices such as magnetic racks.

Features

• This product exhibits strong specificity and high binding capacity to the target protein. Compared to most similar products domestically and internationally, this product has a high antibody binding density, showing strong specificity for proteins with Flag tags. Additionally, the magnetic beads used in this product have a small particle size, reducing non-specific adsorption. Each milliliter of magnetic bead suspension contains approximately 10 mg of magnetic beads and no less than 0.6 mg of Flag antibody. Typically, it can bind to no less than 0.6 mg of Flag-tagged fusion protein, depending on the maximum binding capacity and the molecular weight of the tagged protein. For a sample volume of 500 μL, usually only 10-20 μL of magnetic bead suspension is required to efficiently perform immunoprecipitation experiments.
• This product can bind to various forms of Flag-tagged proteins, including methionine-flag-protein (Met-Flag-Protein) with an N-terminal Flag fusion, N-terminal Flag fusion protein (Flag-Protein), and C-terminal Flag fusion protein (Protein-Flag).
• The binding of the target protein is rapid with this product. The use of nano-sized magnetic beads (~200 nm) provides a large surface area, facilitating rapid and efficient binding between antibodies and antigens. Typically, the antigen absorption process is completed within 10 minutes, and the immunoprecipitation of the target protein is accomplished within 30 minutes. Shortening the operation time effectively prevents degradation or denaturation of the target protein during prolonged procedures, ensuring the preservation of its activity.
• Multiple elution methods can be selected with this product. Depending on the structure of the target protein, its biological function, and the requirements of subsequent applications, various elution methods can be employed, including Flag peptide, acidic elution buffers, and SDS-PAGE sample loading buffers. In particular, elution with Flag peptide eliminates interference from antibody light and heavy chains in Western blot experiments after immunoprecipitation.
• This product is convenient to use. It is stored in a special protective solution that does not contain glycerol. It can be rapidly and efficiently separated using magnetic adsorption, eliminating the need for centrifugation.

Main Indicators

• Product content:    10mg/ml magnetic beads in specific protective buffer
• Beads size:    ~200nm
• Magnetization:    Superparamagnetic
• Coupled antibody:    Anti-Flag mouse monoclonal antibody
• Isotype:    IgG2b
• M.W. of antibody:    Approximately 150kDa
• Antibody concentration:    ≥0.6mg Flag antibody per ml beads
• Binding capacity:    ≥0.6mg Flag‐tagged fusion protein per ml beads
• Specificity:    Met-Flag-Protein, Flag-Protein, Protein-Flag
• Elution method:    Acid, peptide competitive or SDS‐PAGE loading buffer elution.Note: If elute with SDS-PAGE loading buffer, the light (~25kDa) and heavy (~50kDa) chain of Flag antibody will be denatured and release from the beads.
• Application:    IP, Co-IP, Protein purification

Precautions

• This product has been tested and can withstand three or more freeze-thaw cycles without affecting its performance.
• To maintain the pH of the product, it should be kept between 6-8. Avoid high-speed centrifugation, drying, or freezing. Do not leave the magnetic beads in a magnetic field for an extended period as it may cause bead clustering.
• Before use, it is necessary to properly resuspend the product by gently inverting it several times to ensure thorough mixing. Avoid vigorous vortexing or shaking, as it may cause antibody denaturation.
• For immunoprecipitation or purification, it is recommended to include positive and negative control groups.
• After collecting protein samples, it is advisable to proceed with purification as soon as possible and store them at 4°C or on ice to slow down protein degradation or denaturation. To effectively inhibit protein degradation, an appropriate amount of protease inhibitor cocktail can be added to the protein samples.
• If using a vacuum pump or similar instrument to aspirate the supernatant, be cautious of the suction strength to avoid aspirating aggregated magnetic beads.
• Aggregation of the magnetic beads during acidic elution is a normal phenomenon and does not affect their normal use. Adding 0.1% non-ionic detergent (such as Triton X-100, Tween-20, or NP-40) can effectively prevent bead aggregation without affecting antibody binding efficiency.
• High concentrations of DTT, mercaptoethanol, guanidine hydrochloride, or similar substances may have some impact on the binding of this product to the tagged protein.
• This product is intended for scientific research by professionals only and should not be used for clinical diagnosis or treatment, food, or drugs. It should not be stored in regular residential areas.
• For your safety and health, please wear appropriate lab attire and disposable gloves when handling this product.

Storage

Store at 4°C, valid for two years. If not used for an extended period, it can be stored at -20°C, which allows for longer-term storage.

Only for research and not intended for treatment of humans or animals

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