ChromoBlog

How to immunoprecipitate Flag®-tagged proteins

Posted by Christoph Eckert on Dec 16, 2021 10:33:31 AM

Flag®-tag (or DYKDDDDK-tag) is a commonly used short peptide tag for multiple applications such as immunoprecipitation (IP), protein purification, immunofluorescence, and Western blotting (WB). In this blog, we provide an introduction to the IP of Flag®-tagged proteins from cellular extracts. Using our DYKDDDDK Fab-Trap™ as an example, we elaborate on the different steps of IP and highlight the controls that help you achieve the best result in your experiment.

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Topics: Immunoprecipitation

Why is the Fab-Trap™ so Fab-ulous?

Posted by Dr. Bennet Reiter on Nov 25, 2021 9:11:20 AM

Different antibody formats can be used in IP applications, including full-length antibodies, Fab-fragments, and Nanobodies. ChromoTek’s Fab-Trap™ is an IP reagent based on an antibody Fab-fragment. In this blog, we elaborate on the basic principle and advantages of Fab-fragment based reagents for IP. More simply, we ask “Why is the Fab-Trap™ Fab-ulous?”

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Topics: Nano-Traps, Immunoprecipitation, Fab-Trap

Advantages and limitations of different antibody formats in immunoprecipitation

Posted by Dr. Bennet Reiter on Nov 25, 2021 8:52:38 AM

Immunoprecipitation (IP) is a technique used to isolate a protein from a cellular extract. The protein of interest (POI) is recognized by a specific antibody (Ab) conjugated to beads. Different antibody formats can be used in IP, e.g., a full-length Ab, a Fab-fragment, or a Nanobody. Below, we discuss the advantages and limitations of the different Ab formats.

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Topics: Nano-Traps, Immunoprecipitation, Nanobody, Fab-Trap

Co-immunoprecipitation troubleshooting

Posted by Dr. Astrid Sitte on Aug 27, 2021 5:21:16 PM

Co-immunoprecipitation (Co-IP) describes the isolation of a protein and its binding partners from a cell extract using a Nanobody or antibody that is bound to beads. The protein, that directly interacts with the Nanobody, or antibody beads is called “bait”. The binding partner that is indirectly precipitated is called “prey”.

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Topics: GFP-Trap, Immunoprecipitation, Co-IP

How to conduct a Co-immunoprecipitation (Co-IP)?

Posted by Dr. Astrid Sitte on Jul 30, 2021 2:39:52 PM

Co-IP describes the immunoprecipitation of a protein of interest and its interacting partners.

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Topics: GFP-Trap, Immunoprecipitation, Co-IP

Seizing the day: How to save time on your immunoassay experiments

Posted by Dr. Rebecca Northeast (Proteintech) on Feb 3, 2021 10:59:58 AM

When time in the lab is limited, learn how to save precious hours on your immunoprecipitation, immunofluorescence and western blotting experiments.

Due to the current situation with the COVID-19 pandemic, many researchers around the world are faced with restrictions on time and resources in the lab. As we adapt to these changes, the pressure is on to find ways to save time without compromising on results. Proteintech and ChromoTek offer solutions to save you hours on your current immunoassay protocols, helping you achieve publication worthy results in a fast and reliable manner.

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Topics: Immunoprecipitation, Immunofluorescence, GFP, Nano-Secondaries

V5 Tag: Properties, origin, specifications, antibodies & Nanobodies

Posted by Christoph Eckert on Sep 23, 2020 2:09:32 PM

V5 tag is a short peptide tag for detection and purification of proteins. The V5 tag can be fused/cloned to a recombinant protein and detected in ELISA, flow cytometry, immunoprecipitation, immunofluorescence, and Western blotting with antibodies and Nanobodies.

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Topics: Immunoprecipitation, V5

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
  • Magnetic Particles M-270

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
  • Magnetic Particles M-270 for IP of very large proteins/complexes
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Topics: GFP-Trap, Immunoprecipitation, RFP-Trap, GFP Immunoprecipitation, GFP Nanobody, Spot-Trap

What is antibody specificity and affinity? And why is it important for immunoprecipitation?

Posted by Dr. Astrid Sitte on Mar 12, 2020 5:17:58 PM

Introduction of Affinity vs. Specificity

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Topics: Immunoprecipitation

How to obtain a low background in immunoprecipitation assays

Posted by Dr. Astrid Sitte on Mar 10, 2020 2:02:57 PM

Introduction

A high background from unspecific binding of proteins is a common problem in immunoprecipitation (IP). In this blog, the origin of background and different optimization strategies are discussed. Furthermore, it is shown how you can achieve a low background when using the ChromoTek GFP-Trap®, a ready to use reagent for IP of a GFP-tagged proteins.

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Topics: GFP-Trap, Immunoprecipitation

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