RFP-Trap

Schematic pull-down of RFP from human cells transiently expressing RFP. Input (I), non-bound (FT) and bound (B) fractions were separated by SDS-PAGE followed by Coomassie staining and Western Blotting. The bound fraction of the conventional RFP antibodies shows in addition to RFP contaminating heavy chain and light chain. The bound fraction of the RFP-Trap® does not contain these contaminating polypeptide chains.

Description

The ChromoTek RFP-Trap^® is a ready to use affinity resin for immunoprecipitation (IP) of red fluorescent-fusion proteins like mCherry.

RFP-Trap consists of an anti-RFP Nanobody/ V_HH coupled to agarose beads, magnetic agarose beads or Magnetic Beads. ChromoTek’s RFP-Trap is referenced in more than 100 scientific articles.

Specificity
mCherry, mRFP, mRFPruby, mPlum, tagRFP, mKate2 and many more RFP derivatives, see:
Fluorescent protein specificity table

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

Formats/matrices
RFP-Trap Agarose
RFP-Trap Magnetic Agarose
RFP-Trap Magnetic Beads
RFP V_HH, recombinant binding protein
iST RFP-Trap Kit for IP plus sample preparation for MS

Synonyms
anti-RFP V_HH, RFP binding protein, RFP Nanobody, or anti-RFP single domain antibody fragment (sdAb)

Benefit from RFP-Trap for Immunoprecipitation (IP)

Reliable and robust pull-down of mCherry and other RFP-fusion proteins
From cell to gel in less than one hour
High affinity
Strong binding even under harsh washing conditions
No heavy & light chains in your downstream application

What RFP-Trap shall I use?

RFP-Trap Agarose, when lowest background and high binding capacity IP is needed.
RFP-Trap Magnetic Agarose, when magnetic separation and high binding capacity IP is needed.
RFP-Trap Magnetic Beads, when very large proteins/complexes are investigated, and magnetic separation is needed for IP.
iST RFP-Trap kit for immunoprecipitation plus sample preparation for mass spectrometry (MS)

Comparison of matrices & properties

We offer RFP-Trap as Agarose, Magnetic Agarose, and Dynabeads.

	RFP-Trap Agarose	RFP-Trap Magnetic Agarose	RFP-Trap Magnetic Beads
Low background	+++	++	++
Binding capacity	+++	+++	+
Size RFP-tagged protein*	Small to large	Small to large	Small to very large
Bead separation	Centrifugation	Magnetic	Magnetic

* Does depend on protein size and shape, protein multimers, complexes and interaction partners

Extraordinary stable & reliable binding

Dissociation constant K_D of 5 nM
Compatible to harsh wash conditions: 4 M urea, 2 M NaCl, 10 mM DTT, 2 % Nonidet P40 Substitute, 1 % Triton X-100 (RFP-Trap Magnetic Beads: 10mM DTT, 500 mM NaCl, 2 % Nonidet P40 Substitute, 2 % Triton X-100)
Fully applications validated and characterized

Specificity

The RFP-Trap specifically binds to

mCherry, mOrange
mRFP, mRFPruby, mPlum, tagRFP, mKate2, mScarlet

See full fluorescent protein specificity list here.

About RFP

The first red fluorescent protein (RFP) that became commercially available is DsRed. It was isolated from Discosoma sp in 1999. Additional RFPs have been isolated from Anthozoa. Site directed mutagenesis was used to create RFP derivatives with fluorescence in orange, red, and far red colors. Also, monomerized versions were generated and brightness, maturation efficiency, and photostability were improved, making theses RFPs valuable research tools.