NF-κB (p65) Archetype Agency Appraisal is a non-radioactive, acute adjustment for audition specific archetype agency DNA bounden action in nuclear extracts and accomplished corpuscle lysates. A 96-well enzyme-linked immunosorbent appraisal (ELISA) replaces the bulky radioactive electrophoretic advancement about-face appraisal (EMSA). A specific bifold abandoned DNA (dsDNA) arrangement absolute the NF-κB acknowledgment aspect is anchored to the wells of a 96 able-bodied plate. NF-κB independent in a nuclear extract, binds accurately to the NF-κB acknowledgment element. NF-κB (p65) is detected by accession of specific primary antibiotic directed adjoin NF-κB (p65). A accessory antibiotic conjugated to HRP is added to accommodate a acute colorimetric readout at 450 nm. Cayman’s NF-κB (p65) Archetype Agency Appraisal detects animal NF-κB (p65). It will not cross-react with NF-κB (p50).
NF-κB (p65) Archetype Agency Appraisal (Colorimetric) Test Principle The Non-Radioactive NFkB p50/p65 Archetype Agency Appraisal kit is provided in a 96-well format. During the assay, the Capture Probe, a bifold abandoned biotinylated oligonucleotide absolute the accord arrangement for NFkB bounden (5-GGGACTTTCC-3), is alloyed with cellular (nuclear) abstract in the Archetype Agency Appraisal Buffer provided. When incubated together, the alive anatomy of NFkB independent in the nuclear abstract binds to its accord sequence. After incubation, the extract/probe/buffer admixture is again anon transferred to the streptavidin coated plate. The biotinylated bifold abandoned oligonucleotide apprenticed by alive NFkB protein is anchored and any inactive, absolved actual is done away. The apprenticed NFkB archetype agency subunits, p50 and p65, are detected with specific primary antibodies, a Rabbit anti-NFkB p50 (active form) and a Rabbit anti-NFkB p65. An HRP-conjugated accessory antibiotic is again acclimated for apprehension and provides acute colorimetric apprehension that can be apprehend in a spectrophotometric bowl reader. Included in the kit are absolute corpuscle extract, a non-specific bifold abandoned oligonucleotide, and a specific adversary bifold abandoned oligonucleotide. The NF kappa B Archetype Agency Assays were QC activated application nuclear extracts from animal (HeLa) cells. However, because of the attention in the NFkB DNA bounden website and the actuality that the antibodies independent in the kit cantankerous acknowledge with rat and abrasion NFkB, the kit is accepted to plan with samples from rat and abrasion as able-bodied as human. For Research Use Only; Not for use in analytic procedures Use of this appraisal in NFkB-related biologic analysis may be covered beneath U.S. Patent No. 6,150,090 and crave a authorization from Ariad Pharmaceuticals (Cambridge, MA, USA).
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