V5-Trap Magnetic Agarose

Description
The ChromoTek V5-TrapTM Magnetic Agarose is an affinity resin for immunoprecipitation of V5-tagged proteins.
It consists of a anti V5 Nanobody/VHH coupled to magnetic agarose beads.

Specificity
V5-tag amino acid sequence GKPIPNPLLGLDST at the N-terminus, C-terminus, or internal site of the fusion protein.

Applications
Immunoprecipitation/ Co-IP
Mass spectrometry
ChIP/ RIP analysis
On-bead enzyme assays

Product Size Code Price Buy
Product V5-Trap Magnetic Agarose Size 250 µL (10 reactions) Code v5tma-10 Price $ 275
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Product V5-Trap Magnetic Agarose Size 500 µL (20 reactions) Code v5tma-20 Price $ 475
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Product V5-Trap Magnetic Agarose Size 2.5 mL (100 reactions) Code v5tma-100 Price $ 2050
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Product V5-Trap Magnetic Agarose Size 5 mL (200 reactions) Code v5tma-200 Price Please inquire
Product V5-Trap Magnetic Agarose Size 10 mL (400 reactions) Code v5tma-400 Price Please inquire
Product V5-Trap Magnetic Agarose Kit Size 500 µL (20 reactions) Code v5tmak-20 Price $ 595
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Product V5-peptide Size 1 mg Code v5p-1 Price $ 85
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Product Binding Control Magnetic Agarose Beads Size 500 µL (20 reactions) Code bmab-20 Price $ 75
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Product V5 VHH, recombinant binding protein Size 250 µL Code v5t-250 Price Please inquire

Affinity
Dissociation constant KD of 40 nM

Wash buffer compatibility
2 % Nonidet P40 Substitute, 2 % Triton X-100, 1 M NaCl, 5 mM TCEP, 5 mM DTT, 10 mM ß-mercaptoethanol, 4 M Urea

Specificity
V5-tag sequence motif GKPIPNPLLGLDST

Binding capacity
10 µL slurry bind ~ 7 µg of recombinant V5-tagged protein ~30 kDa

Coupled Nanobody/ VHH
recombinant, monoclonal anti-V5-tag single domain antibody (sdAb) fragment

Bead properties
Bead size: ~ 40 µm
Storage buffer: 20% EtOH

Elution
- V5-peptide. For details see below.
- SDS sample buffer
- 0.1 M glycine pH 2.0
Please consider whether elution is really needed. We recommend on-bead assays like on-bead digestion for MS analysis.

Compatibility with mass spectrometry
The V5-Trap is optimized for on-bead digestion (see protocol). 

How to cite this product
RRID:  AB_2868498

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.
Please find more information here:

Preomics kit

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 traps is very high.

Benefits

  • No heavy & light antibody chains
  • Pull-down of V5-tagged proteins
  • Harsh washing conditions
  • Elution of native proteins
  • One step immunoprecipitation
  • Reproducible results

V5-Trap Magnetic Agarose Kit

The V5-Trap Magnetic Agarose is also available in a kit, including:

  • V5-Trap Magnetic Agarose
  • Lysis*, dilution and wash buffers

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

Binding Control for Magnetic Agarose Beads

V5-peptide

The V5-peptide may be used to elute native and functional V5 fusion proteins:

V5-peptide for effective elution of V5 fusion proteins

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