Description
The TransMax™ kits are chemiluminescence-based assays that enable rapid
analysis of transcription factor activity in a convenient 96-well format. These
sensitive, non-radioactive assays can be completed within four hours and are
a superior alternative to standard gel shift assays, immunoblotting, and antibodies
based ELISAs. The TransMax kits employ a proprietary assay chemistry and chemiluminescent
detection and offer superior sensitivity, broad dynamic range, and excellent
reproducibility compared to the standard assay approaches.
Highly Sensitive
Assays Yielding Specific Results |
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| NFκB binding activity was detected in as little as
0.004 gel shift units (GSU) of recombinant p50. (1 GSU = amount of purified
NFκB protein required to completely shift 380 fmoles of NFκB
binding site in a standard gel shift assay.) |
TransMax EGR Kit detects EGR activation in 2.5 µg
of nuclear extract from Jurkat cells treated with TPA for 4 hours. Excess
of unlabeled EGR binding site reduced the signal by more than 90%. Mutants
binding sites predicted not to bind EGR did not reduce the specific assay
signals. |
The TransMax kits utilize an elegant direct quantification approach to measure
the binding of activated transcription factors to their nascent DNA binding
sequences. First, nuclear extracts from cells or tissues are incubated with
a biotin-labeled DNA probe consisting of a double-stranded consensus transcription
factor binding site and a single -stranded capture region (Figure 1). Activated
transcription factors bind specifically to the double-stranded consensus sequence.
Next, a double-stranded DNA specific nuclease is added to the mixture. Probes
with a bound transcription factor are protected from digestion whereas unbound
probes are degraded.

The reactions are then transferred to a 96-well plate coated with the complementary
strand to the capture region of the probe, and the intact probe/transcription
factor is captured on the plate. After unbound biotin and digested probes are
washed away, the intact captured biotinylated probes are detected using a streptavidin-alkaline
phosphatase conjugate and a chemiluminescent substrate. The amount of signal
generated is proportional to the amount of transcription factor in the nuclear
extract and can be easily detected using any standard microplate luminometer.
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| Detection of NFκB in small samples.
NFκB activation was observed in as little as 0.1 µg of nuclear
extract from TNF-a stimulated HeLa cells. |
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| Detection of Multiple Transcription Factor Complexes.
TransMax AP-1 Kit detects both c-jun homodimers and c-jun-c-fos heterdimers,
obviating the need for specific antibodies to multiple protein forms or
splice variants. |
Contents
Each TransMax kit comes with all the reagents you need to quantitatively measure
transcription factors including one PCR plate, one capture plate, two plate
sealers, sample diluents, binding mixes, digestion reagents, hybridization buffer,
10X washing buffer, conjugation reagents, and chemilumiinescent substrate.
Storage
Store all components at -20ºC upon receipt. Stable for 6 months when stored proprely.
Once diluted the Wash buffer may be stored at room temperature.