mNeonGreen-Trap Magnetic Agarose
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I: Input, FT: Flow-through, B: Bound
Description
The ChromoTek mNeonGreen-Trap Magnetic Agarose is an affinity resin for immunoprecipitation of mNeonGreen-fusion proteins.
It consists of a mNeonGreen Nanobody/ VHH coupled to magnetic agarose beads.
Specificity
mNeonGreen, Branchiostoma lanceolatum
Applications
Immunoprecipitation/ Co-IP
Mass spectrometry
On-bead enzyme assays
ChIP/ RIP analysis
| Product | Size | Code | Price | Buy |
|---|---|---|---|---|
| Product mNeonGreen-Trap Magnetic Agarose | Size 250 µL | Code ntma-10 | Price $ 275 |
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| Product mNeonGreen-Trap Magnetic Agarose | Size 500 µL (20 reactions) | Code ntma-20 | Price $ 475 |
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| Product mNeonGreen-Trap Magnetic Agarose | Size 2.5 mL (100 reactions) | Code ntma-100 | Price $ 2050 |
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| Product mNeonGreen-Trap Magnetic Agarose | Size 5 mL (200 reactions) | Code ntma-200 | Price Please inquire | |
| Product mNeonGreen-Trap Magnetic Agarose | Size 10 mL (400 reactions) | Code ntma-400 | Price Please inquire | |
| Product mNeonGreen-Trap Magnetic Agarose Kit | Size 500 µL (20 reactions) | Code ntmak-20 | Price $ 595 |
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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 mNeonGreen-Binding Protein | Size 250 µL | Code nt-250 | Price $ 275 |
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Affinity
Dissociation constant KD of 2 nM
Wash buffer compatibility
4 M urea, 2 M NaCl, 10 mM DTT, 2 % Nonidet P40 Substitute, 1 % Triton X-100, 0.2 % SDS
Specificity
mNeonGreen, Branchiostoma lanceolatum
Binding capacity
10 µL slurry bind more than 12 µg of recombinant mNeonGreen
Coupled Nanobody/ VHH
recombinant, monoclonal anti-mNeonGreen single domain antibody (sdAb) fragment
Bead properties
Bead size: ~ 40 µm
Storage buffer: 20 % EtOH
Elution
- 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 mNeonGreen-Trap is optimized for on-bead digestion. Complete tryptic digest results in 5-6 peptides.
Storage instructions
Shipped at ambient temperature. Upon receipt store at 4°C; stable for one year.
Protocol
Brochure
Trouble shooting
SDS
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 can I elute bound proteins from a trap in their native state?
You can elute your fusion protein of interest with 0.2 M glycine pH 2.5 at room temperature. Pipette the beads up and down for 60-120 seconds and repeat this step. Ensure to neutralize your supernatant immediately afterwards by adding 1 M Tris base pH 10.4.
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:
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
- Superior pulldown of mNeonGreen-fusion proteins
- No heavy & light chains in your downstream application
- Strong binding even under harsh washing conditions
- Short incubation time of about 60 minutes
mNeonGreen-Trap Magnetic Agarose Kit
The mNeonGreen-Trap Magnetic Agarose is also available in a kit, including:
- mNeonGreen-Trap Magnetic Agarose
- Lysis*, wash and elution 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
- Control for unspecific binding of proteins, DNA, etc. to beads
- Pre-clearing of cell lysate
- Binding control for mNeonGreen-Trap Magnetic Agarose
Only for research applications, not for diagnostic or therapeutic use!