Description
Overview
MAb Anti cmyc-Tb is an IgG1 raised against a synthetic peptide corresponding to the 408-439 sequence of human c-myc protein labeled with Tb. It recognizes the EQKLISEEDL motif and is specific for human c-myc, although a slight cross-reaction has been observed with murine c-myc at high antibody concentration.
This reagent can be used in both biochemical and cellular formats to study a wide variety of interactions: protein/protein, protein/peptide, protein/DNA, receptor/ligand.
HTRF can detect a broad range of affinity constants ranging from picomolar to low millimolar.
HTRF assays offer many advantages over other technologies:
Homogeneous add-and-read format
No wash steps
Low background
Straightforward miniaturization from 96- or 384-well microplates to high density assay formats such as 384-well low volume and 1536-well plates
Stable signal, providing flexibility in time of readout or size of assays
How it works
Assay principle
In an HTRF interaction assay, one partner is labeled (directly or indirectly) with the donor, and the other with the acceptor (again, directly or indirectly). The intensity of the signal is proportional to the binding of the 2 partners. In the example shown here: MAb Anti cmyc-Eu cryptate binds to the c-Myc tagged partner A, while partner B* binds to a specific Ab labeled with an HTRF acceptor. *partner B can also be biotinylated, tagged, or Fc fused. In these cases, use the corresponding HTRF reagent (anti-Tag, anti-species, protA, Streptavidin) labeled with acceptor for the detection.
Assay protocol
The example on the right describes the protocol using a 20 µL final assay volume for detecting an interaction between a c-Myc-tagged partner A and a non-tagged partner B*. Dispense the 2 partners (10 µL), incubate, add MAb Anti cmyc-Eu cryptate (5 µL) and anti-partner B labeled with acceptor (5 µL), incubate, and read. *partner B can also be biotinylated, tagged, Fc fused or directly labeled. In these cases, use the corresponding HTRF reagent (anti-Tag, anti species, protA, Streptavidin) labeled with acceptor for the detection.
