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TransAM® Elk-1

dna-binding ELISA for activated Elk-1 transcription factor

TransAM® Kits are DNA-binding ELISAs that facilitate the study of transcription factor activation in mammalian tissue and cell extracts. Assays are available for over 40 different targets (see the list at right). Each kit includes a 96-stripwell plate in which multiple copies of a specific double-stranded oligonucleotide have been immobilized. When nuclear or whole-cell extract is added, activated transcription factor of interest binds the oligonucleotide at its consensus binding site and is quantified using the included antibody, which is specific for the bound, active form of the transcription factor being studied. For complete details, click the TransAM® Method tab below.

TransAM® Elk-1 Transcription Factor ELISA Kits

TransAM Elk-1 Kits provide everything needed to study activated Elk-1, member of ETS oncogene family, including a positive control extract. The Elk-1 kit can be used to detect Elk-1 phosphorylated at Ser383 in human extracts. Click the Elk1 Info tab below for data and more information; kit manuals can be downloaded under the Documents  tab.

Name Format Cat No. Price  
TransAM® Elk-1 1 x 96 rxns 44396 $665 Buy Now
5 x 96 rxns 44896 $2,795 Buy Now

Elk1 Transcription Factor Info

Elk-1, member of ETS oncogene family, is a transcription factor that is activated by mitogen-activated protein kinases (MAPKs) and whose main function is the regulation of growth-related proteins in response to extracellular stimuli. Ets family members share a Ets domain and a winged helix-loop-helix DNA binding domain that recognizes a GGAA/T-based sequence. Phosphorylation of Elk-1 increases the likelihood of ternary complex formation and binding with the serum response factor and the serum response element in the c-fos promoter. Elk-1 can be stimulated by the ERK1/2, JNK and p38 MAPK pathways to regulate the activity of the c-fos proto-oncogene in response to extracellular stimuli. Elk-1 is also a nuclear target for the ras-raf-MAPK signaling cascade. TransAM Elk-1 Kits contain antibody directed against Elk-1 phosphorylated at serine 383.

Active Motif also offers our FACE™ In-cell Western Phospho ELISAs for quantifying and comparing the phosphorylated and total amounts of ERK1/2, JNK and p38 proteins.

TransAM Elk-1 data 
Figure 1: Monitoring Elk-1 activity in different cell lines with the TransAM Elk-1 Kit.

Different amounts of nuclear extracts from untreated U-937, untreated HeLa and HeLa (TPA, 10 min) cells are tested for Elk-1 activity using the TransAM Elk-1 Kit.

The TransAM® transcription factor ELISA advantage

Historically, transcription factor studies have been conducted using gelshift, Western blot and reporter plasmid transfections, which are time-consuming, do not allow for high-throughput and provide only semi-quantitative results. TransAM assays are up to 100 times more sensitive than gelshift techniques, and can be completed in less than 5 hours. Because TransAM is an ELISA-based assay*, there is no radioactivity, and the high-throughput stripwell format enables simultaneous screening of 1-96 samples. Inconsistencies due to variable reporter plasmid transfections are eliminated, along with the need to construct stable cell lines.

Why use TransAM® transcription factor ELISAs?

  • Up to 100-fold more sensitive than gelshift assays
  • Eliminates the use of radioactivity and the need to run gels
  • Results in less than five hours
  • Colorimetric readout enables easy, quantitative analysis with spectrophotometry at 450 nm
  • 96-stripwell format enables both high and low throughput

How TransAM® transcription factor ELISAs work

The TransAM format is perfect for assaying transcription factor binding to a consensus-binding site. TransAM Kits contain a 96-stripwell plate to which the consensus-binding site oligo has been immobilized. Activated nuclear extract is added to each well and the transcription factor of interest binds specifically to this bound oligonucleotide. A primary antibody specific for an epitope on the bound and active form of the transcription factor is then added followed by subsequent incubation with secondary antibody and Developing Solution to provide an easily quantified, sensitive colorimetric readout (Figure 1).

Flow chart of the TransAM DNA binding transcription factor ELISA method for measurement of activated transcription factors
Figure 1: Flow chart of the TransAM process.

Activated transcription factor in the cell extract binds to the consensus-binding site on the oligo immobilized in the well. Incubation with the supplied primary and secondary antibodies specifically quantifies the amount of activated transcription factor.

* Technology covered under EAT-filed patents and licensed to Active Motif. Use of TransAM in NFκB-related drug discovery may be covered under U.S. Patent No. 6,150,090 and require a license from Ariad Pharmaceuticals (Cambridge, MA, USA).

Contents & Storage

One or five Elk-1 96-well plate(s) with plate sealer(s), phosphorylated Elk-1 primary antibody, HRP-conjugated secondary antibody, wild-type and mutated oligonucleotides, positive control cell extract, DTT, Poly [d(I-C)], Protease Inhibitor Cocktail, Lysis, Binding, 10X Washing and 10X Antibody Binding Buffers, and Developing and Stop Solutions. Reagent storage conditions vary from room temperature to -80°C, see manual for details. All reagents are guaranteed stable for 6 months when stored properly.