ChromoBlog

Which beads should I use for my immunoprecipitation?

Posted by Dr. Astrid Sitte on Sep 16, 2020 9:42:57 AM

ChromoTek's Nano-Traps are optimized for the immunoprecipitation (IP) of proteins and their interacting factors. Nano-Traps comprise of a Nanobody/ VHH conjugated to beads. For 3 Nano-Traps, e.g. GFP-Trap®RFP-Trap®, and Spot-Trap®, we offer different types of beads:

  • Agarose beads
  • Magnetic Agarose beads
  • Dynabeads™

Immunoprecipitation with GFP-Trap.
I: Input, FT: Flow-Through, B: Bound

Based on the properties of the different beads (see table), we recommend:

  • Agarose beads for very low background and high binding capacity IP
  • Magnetic Agarose beads for magnetic separation and high binding capacity IP
  • Dynabeads for IP of very large proteins/complexes
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Topics: GFP-Trap, Immunoprecipitation, RFP-Trap, GFP Immunoprecipitation, GFP Nanobody, Spot-Trap, Dynabeads

GFP-Trap in multiwell plates

Posted by Christoph Eckert on Mar 19, 2020 2:10:56 PM

GFP-Trap®Multiwell Plate for convenient high throughput analysis of protein interactions. It consists of our anti-GFP VHH immobilized in multiwell plates. The GFP-Trap Multiwell Plate is optimized for immunoprecipitation (IP), IP-MS, and ELISA. Here you can find the protocol for sandwich ELISA und immunoprecipitation (IP) with GFP-Trap

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Topics: GFP Immunoprecipitation

5 Tips for Successful Immunoprecipitations (IP)

Posted by Klaus Herick on Mar 3, 2020 3:15:19 PM

The pull-down of proteins is an established technology. It can be difficult, particularly when the protein of interest is expressed at low levels. Here we discuss 5 tips that help you to improve your IP results.

 

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Topics: Immunoprecipitation, GFP Immunoprecipitation

What GFP-Trap should I use for my immunoprecipitation?

Posted by Dr. Astrid Sitte on Jan 20, 2020 4:42:38 PM

ChromoTek GFP-Trap® is optimized for the immunoprecipitation of GFP-tagged proteins and their interacting factors.

The GFP-Trap consists of the ChromoTek anti-GFP Nanobody/ VHH that is coupled to 3 different types of beads or is immobilized in a 96 multiwell plate:

  • Agarose beads
  • Magnetic Agarose beads
  • Dynabeads™
  • 96 Multiwell Plate

Immunoprecipitation with GFP-Trap.
I: Input, FT: Flow-Through, B: Bound

Based on the properties of the different matrices (see table), we recommend:

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Topics: GFP-Trap, GFP Immunoprecipitation, GFP Nanobody, Dynabeads

GFP-Trap Dynabeads: Most cited GFP-Nanobody conjugated to Dynabeads

Posted by Christoph Eckert on Jan 17, 2020 11:27:30 AM

GFP-Trap® Dynabeads is optimized for the immunoprecipitation (IP) of large GFP-tagged proteins and for Co-IP of protein complexes. It consists of ChromoTek’s established anti-GFP Nanobody conjugated to Dynabeads™. Hence, also GFP-Trap Dynabeads has the very high affinity of 1 pM like GFP-Trap Agarose and Magnetic Agarose.

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Topics: GFP Immunoprecipitation, GFP Nanobody, Dynabeads

Which are 2018's most used Antibodies?

Posted by Klaus Herick on Aug 26, 2019 4:29:03 PM

 
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Topics: GFP-Trap, GFP Immunoprecipitation, GFP VHH, GFP Nanobody

Working with green fluorescent proteins: Tools and properties

Posted by Christoph Eckert on Feb 7, 2019 1:18:14 PM

Jellyfish Green Fluorescent Protein (GFP) and its derivatives are still the most frequently used fluorescent proteins in biomedical research. Recently, additional green fluorescent proteins have been discovered in higher animals such as crustaceans and lancelets. These FPs share a common fold, but diverge widely in their primary sequence. Thus, they require novel, dedicated antibody research tools. Here is an overview about EGFP (the most commonly used GFP derivative), TurboGFP and mNeonGreen.

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Topics: Nanobody, mNeonGreen, mNeonGreen immunoprecipitation, GFP, GFP Immunoprecipitation, TurboGFP, TurboGFP immunoprecipitation, EGFP

On-bead digestion protocol: From immunoprecipitation with nanobodies to MS analysis

Posted by Christoph Eckert on Sep 15, 2017 3:52:48 PM

What makes on-bead digestion favorable?

Just pull down your protein of interest with immobilized nanobodies, also termed VHHs or single domain antibodies. Then follow the on-bead digestion protocol (see below) and submit the digest to your core facility for effective mass spectrometer analysis of (co-) precipitated proteins.

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Topics: GFP-Trap, GFP Immunoprecipitation, GFP VHH, GFP Nanobody, on-bead digestion, GFP mass spec

The best anti-GFP antibody for immunoprecipitation: GFP-Trap

Posted by Christoph Eckert on Sep 7, 2017 1:05:34 PM

Life science laboratories apply green fluorescent proteins (GFP) to study protein localization, interaction and dynamics in fluorescence microscopy. Immunoprecipitation (IP), mass spectrometry (MS), co-immunoprecipitation (Co-IP) and/or affinity purification investigate more aspects including posttranslational modifications (PTMs), DNA binding, and protein-protein interaction. Here, we compare two different antibody systems for immunoprecipitation of GFP-fusion proteins: GFP-Trap and anti-GFP IgG antibody

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Topics: GFP-Trap, GFP, GFP Immunoprecipitation, Best GFP antibody for IP, Best antibody for immunoprecipitation, GFP VHH, GFP Nanobody

How to prepare protein interaction partners for MS analysis

Posted by Christoph Eckert on Apr 24, 2017 11:02:03 AM

Immunoprecipitation (IP) followed by mass spectrometry (MS) analysis is a powerful method to identify interaction partners of a protein of interest. However, sometimes it can be difficult to obtain reliable results.

Now, the combined use of ChromoTek GFP-Trap for immunoprecipitation and PreOmics iST sample preparation kit is in fact a straight forward, reproducible, and reliable approach to identify protein interaction partners. It even preserves posttranslational modification (PTM) depending protein-protein interactions.

In a recent applications note we have demonstrated how protein interaction partners of PARP1 can be identified: We have immunoprecipitated eGFP-tagged PARP1 protein and its interacting partners using the GFP-Trap_M and subsequently processed the pulldown for MS analysis following the instructions of the iST kit.

PARylation mediates the interaction of PARP1 to several DNA repair proteins. Our results show that those interactions are not disrupted by neither the IP nor the MS sample preparation used herein. Therefore, the streamlined combination of the GFP- Trap and the iST kit’s workflow proves to preserve PTM-mediated protein-protein interactions.

 

Download application note

 

The PreOmics iST kit can also be used with other ChromoTek Nano-Traps.

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Topics: GFP-Trap, Immunoprecipitation, Mass spec, GFP Immunoprecipitation, GFP MS

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