NF-KB -- One New Target for Cancer Therapy

NF-κB(Nuclear Factor Kappa B) is a transcription factor, it is activated by various intra- and extra-cellular stimuli such as cytokines, oxidant-free radicals, ultraviolet radiation, and bacterial or viral products. Activated NF-κB enters cell nucleus and promotes biological functional genes widely expressing. Under circumstances, more than 200 genes target NF-κB molecule in various types of cells. NF-κB not only functions in the regulation of immune and inflammatory response, but also in the proliferation, migration, and apoptosis. More and more evidences show that NF-kB plays an important role in neoplasia, and the abnormal expression of NF-kB has been detected in a lot of human cancers. Recently numerous researches have been focused on the influence of NF-kB activation on signal transduction mechanism, so, NF-kB has been the new target for cancer therapy.

The Cloud-Clone Corp. developed a series of detection tools for NF-κB which include recombinant protein, antibodies and ELISA kits. The development of NF-κB related products bring researchers great access to researches on NF-κB.

The rough development processes of NF- κ B related products are shown as below.

At first, the expression plasmid was constructed by PCR amplified NF-κB gene and pUSCNK-1 vector. After transforming and sequencing correctly, the NF-κB expression plasmid was inducible expressed by IPTG. Then it was purified by Nickel column as there’s a his-tag in protein sequence, and recombinant NF-κB with high purity (RPB824Hu01, RPB824Hu02, RPB824Mu01, RPB824Ra01, RPB824Ga01) were obtained. After that, molecular weight and purity were tested by SDS-PAGE (Fig. 2).

Secondly, mice were immunized for monoclonal antibody preparation, after purification, various monoclonal antibodies specific to NF-κB in different species (MAB824Hu22, MAB824Ra21, MAB824Ga21) were obtained.

In the meanwhile, rabbit were immunized for polyclonal antibody preparation, after the antiserum were obtained and purified, polyclonal antibodies specific to NF-κB in different species (PAB824Hu01, PAB824Hu02, PAB824Mu01, PAB824Ra01, PAB824Ga01) were obtained. Then these antibodies were coupled with biotin to prepare PAB824Hu71 and PAB824Mu71, coupled with FITC to prepare PAB824Hu81 and PAB824Mu81. All these antibodies have been tested by Western blot and IHC. The result of PAB824Hu01 used for serum sample detection in WB is shown in Fig 3, and the result of PAB824Hu01 in IHC test is shown in Fig 4. Sample used in IHC test is human heart tissue, and it turns that the protein was mainly expressed in the cytoplasm and nucleus.

For more information, please visit http://www.cloud-clone.us/.