DYKDDDDK Fab-Trap Agarose

Description
The DYKDDDDK Fab-Trap™ Agarose is a premium affinity resin for efficient  immunoprecipitation (IP) of Flag®-tagged proteins. The DYKDDDDK Fab-Trap Agarose consists of a Fab-fragment, which is covalently bound to agarose beads.

Specificity
The DYKDDDDK Fab-Trap™ Agarose specifically binds to 1x and 3xDYKDDDDK-tag sequence (1x and 3xFlag®-tag). It binds to same epitope as Sigma's Anti-FLAG® M2 Antibody and is compatible with N-, C-terminal or internal tagging.

Applications
Immunoprecipitation (IP) / Co-IP
Mass spectrometry (MS)
Protein purification
ChIP/ RIP analysis
On-bead enzyme assays

Try out the DYKDDDDK IP Starter Pack (ffap)
DYKDDDDK Fab-Trap™ Agarose (20 reactions, product code ffa-20) for immunoprecipitation, 3xDYKDDDDK-peptide (1mg, fp-1) for elution and DYKDDDDK tag Polyclonal antibody (Binds to FLAG® tag epitope) (20 µL, 20543-1-AP-20UL) for detection in Western blots.

Product Size Code Price Buy
Product DYKDDDDK Fab-Trap Agarose Size 250 µL (10 reactions) Code ffa-10 Price $ 275
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Product DYKDDDDK Fab-Trap Agarose Size 500 µL (20 reactions) Code ffa-20 Price $ 475
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Product DYKDDDDK Fab-Trap Agarose Size 2.5 mL (100 reactions) Code ffa-100 Price $ 2050
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Product DYKDDDDK Fab-Trap Agarose Size 5 mL (200 reactions) Code ffa-200 Price Please inquire
Product DYKDDDDK Fab-Trap Agarose Size 10 mL (400 reactions) Code ffa-400 Price Please inquire
Product DYKDDDDK Fab-Trap Agarose Kit Size 500 µL (20 reactions) Code ffak-20 Price $ 595
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Product 3xDYKDDDDK-peptide Size 1 mg Code fp-1 Price $ 105
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Product DYKDDDDK Immunoprecipitation Starter Pack Size 20 reactions Code ffap Price $ 540
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Product Binding Control Agarose Beads Size 500 µL (20 reactions) Code bab-20 Price $ 75
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Product Spin Columns Size 10 units Code sct-10 Price $ 35
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Product Spin Columns Size 20 units Code sct-20 Price $ 60
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Product Spin Columns Size 50 units Code sct-50 Price $ 130
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Wash buffer compatibility
0.5 % Nonidet P40 Substitute, 5 % Tween™ 20, 2 M NaCl, 10 mM TCEP, 10 mM DTT, 10 mM ß-mercaptoethanol, 2 M Urea

Specificity
1x and 3xDYKDDDDK-tag sequence (1x and 3xFlag®-tag). The DYKDDDDK Fab-Trap™ Agarose binds to same epitope as Sigma's Anti-FLAG® M2 antibody.

Binding capacity
25 µL slurry bind about 12.5 – 15 µg of recombinant 3xDYKDDDDK-tagged protein (~30 kDa)

Clone
Recombinant Fab fragment of monoclonal mouse IgG1 M2.1

Bead properties
Bead size: ~ 90 µm (cross-linked 4 % agarose beads)
Storage buffer: 1x PBS, Preservative: 0.09 % sodium azide

Elution
- 3xDYKDDDDK-peptide
- SDS sample buffer
- 0.2 M glycine pH 2.5
Please consider whether elution is really needed. We recommend on-bead assays like on-bead digestion for MS analysis.

Compatibility with mass spectrometry
The DYKDDDDK Fab-Trap Agarose is optimized for on-bead digestion. 

How to cite this product
RRID: AB_2894836

Storage instructions
Shipped at ambient temperature. Upon receipt store at 4°C. Stable for one year.

Should I use an N-terminal or C-terminal fusion tag?

Both the N-terminal or C-terminal fusion tag work well with the traps.

How can I avoid unspecific protein interactions binding to the trap?

For preclearing of your sample we recommend to use our binding controls (bab-20 or bmab-20).

Please find more information in our Troubleshooting guide

How many mammalian cells are required for an immunoprecipitation reaction?

For one immunoprecipitation reaction, we recommend using ~10^6 - 10^7 mammalian cells. The yield is also dependent on the expression level of your protein of interest and the interaction partners.

How much cell extract should I use for an immunoprecipitation reaction?

For other type of cells than mammalian cells, we recommend using 0.5 - 1.0 mg of cell extract.

Do I need to elute bound proteins from the beads for mass spectrometry analysis?

No, you can directly conduct an on-bead digestion after immunoprecipitation. This procedure allows faster and more efficient sample preparation and a potential higher yield.

Protocol on-bead digestion

Do I need to elute my protein of interest from the beads for enzymatic assays?

No, you can directly perform your enzymatic assay on the beads if the active center is not blocked.

Please find more information in our application note "enzymatic activity assay"

What is the amount of trap slurry I need for one immunoprecipitation reaction?

25 µL slurry are sufficient for one pull-down reaction as the affinity of the DYKDDDDK Fab-Trap is very high.

DYKDDDDK Immunoprecipitation Starter Pack

You look for a cost-effective pack for Flag® IP or you are planning your first IP of a Flag-tagged protein? Or you want to test the DYKDDDDK Fab-Trap™ Agarose?

Here is a Starter Pack (ffap) containing

Benefits

  • Pulldown of Flag®-tagged proteins
  • Lowest background in class
  • Less contaminating antibody chains
  • Reliable: recombinantly expressed Fab-fragment derived from M2 clone
  • Small Fab-fragment tolerates harsh wash buffer conditions
  • Convenient and fast: Easy handling and ready-to-use beads
  • Highly selective binding of both N-and C-terminal Flag-tag

3xDYKDDDDK-peptide

The 3xDYKDDDDK-peptide may be used for the elution of native and functional Flag-tagged proteins bound to DYKDDDDK Fab-Trap or anti-DYKDDDDK antibodies.

The 3xDYKDDDDK-peptide  has the amino acid sequence DYKDDDDK DYKDDDDK DYKDDDDK.

DYKDDDDK Fab-Trap Agarose Kit

The DYKDDDDK Fab-Trap Agarose is also available in a kit, including:

  • DYKDDDDK Fab-Trap Agarose
  • Lysis*, dilution and wash buffers

*The lysis buffer provided is suitable for mammalian cells. For other cell types use another suitable lysis buffer.

Spin columns

  • Easy and convenient handling
  • Rapid washing and clean elution of bound proteins
  • Simplify the pulldown of your protein
  • Spin columns for DYKDDDDK Fab-Trap Agarose

Binding Control for Agarose Beads

  • Control for unspecific binding of proteins, DNA, etc. to beads
  • Pre-clearing of cell lysate
  • Binding control for DYKDDDDK Fab-Trap Agarose

Why is the DYKDDDDK Fab-Trap superior?

  1. Minimal contamination by Fab-fragment of the DYKDDDDK Fab-Trap
    The DYKDDDDK Fab-Trap consists of a Fab-fragment, which is covalently bound to beads. In contrast to antibody-based immunoprecipitation reagents, there is no contamination from DYKDDDDK Fab-Trap on your SDS-PAGE and Western blot above 25 kDa, resulting in cleaner SDS-PAGE and Western blots. Moreover, you will find less antibody derived peptide contaminations in subsequent mass spectrometric analysis.
  2. Lowest Background in class
    ChromoTek’s DYKDDDDK Fab-Trap is an anti-Flag affinity reagent based on a Fab-fragment, which is only a third of the size of a conventional antibody. Due to its smaller size, the Fab-fragment minimizes unwanted interaction with the host-cell proteome, resulting in efficient immunoprecipitations with superior purity and less background.
  3. Small Fab-fragment tolerates harsh wash buffer conditions
    The small Fab-fragment of the DYKDDDDK Fab-Trap tolerates harsher washing conditions as comparable anti-Flag antibody affinity resins. The possibility to use harsher wash buffers, enables effective removal of unwanted cellular proteins for highest purity immunoprecipitation.
  4. Tight and highly selective binding of a single Flag-tag
    The DYKDDDDK Fab-Trap binds with high specificity to N- or C-terminal Flag-tagged proteins. In addition, it has a high affinity to both 1x or 3xFlag-tag enabling a broad experimental applicability.
  5. Ready-to-use reagent
    DYKDDDDK Fab-Trap is a ready-to-use reagent that comes pre-conjugated to beads. There is no need for tedious and time-consuming pre-incubation steps required for pulldowns with protein A or G beads making the immunoprecipitation faster, more convenient, and easier to reproduce.
  6. Mild, competitive elution with DYKDDDDK-peptide or acidic conditions
    DYKDDDDK Fab-Trap can be mildly eluted with 3xDYKDDDDK-peptide, preserving the native structure of your Flag-tagged protein for structural and activity downstream analysis. In addition, convenient elution by low pH or SDS-sample buffer offers fast and efficient elution performance.
  7. Reproducible results
    DYKDDDDK Fab-Trap binds to the same epitope as Sigma’s ANTI-FLAG® M2 antibody. The Fab-fragment of the DYKDDDDK Fab-Trap is recombinantly produced for the generation of highly reliable data.

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Only for research applications, not for diagnostic or therapeutic use!