DNA-binding ELISA kits that semi-quantitatively detect active transcription factors — NF-κB, p53, CREB, SMAD, STAT, c-Jun, Myc and many more — directly from nuclear or cell lysates, with no radioactivity and no gel shift.
How the assay detects active transcription factors
The Transcription Factor Activity Assay is a qualitative indirect ELISA. Streptavidin is bound to the plate, and biotinylated double-stranded consensus oligonucleotides are captured via the high-affinity biotin–streptavidin interaction. After blocking, the sample is added and the active transcription factor binds its DNA motif.
A primary antibody binds the DNA-bound factor, an HRP-conjugated secondary antibody is added, and TMB substrate produces a blue product that turns yellow on stopping. Absorbance at 450 nm is proportional to the amount of active transcription factor in the sample.
FAQs
Transcription factor assay FAQs
My kit was left at room temperature for several days — can I still use it?
It is most likely still usable, though activity may be slightly reduced. The most temperature-sensitive components are the standards, HRP-streptavidin and detection antibody.
Why am I seeing a weak or no signal?
Common causes include an unsuitable nuclear lysate, poor lysate preparation or storage, or missed steps. Add protease and phosphatase inhibitors, keep everything on ice, store at −80°C avoiding freeze-thaw, and follow the manual closely for reagent volumes and storage.
Why is there high background or uneven colour development?
Usually inadequate or uneven washing, too much antibody, lysate or substrate, over-long substrate incubation, or contamination. Wash thoroughly with the correct volumes, reduce antibody/lysate/substrate as needed, use sterile technique, and ensure your multichannel pipette or plate washer is calibrated.
Don’t see your transcription factor?
We cover 100+ transcription factors across total-activity and phospho-specific formats, and can develop custom DNA-binding assays on request.
