Preparation Of Two Polyclonal Antibodies Of ILK And Their Primary Application

ILK (Integrin-linked kinase) is an important Ser/Thr protein kinase in cytoplast ,which acts as the effector of integrin, growth factor and Wnt signaling pathway, mediates many biological actions such as cell adhesion and signal transduction, plays a critical role in cell invasiveness and metastasis. Recent study suggested that ILK over-expression or constitutive activation may regulate the progression of cellular abnormalities from benign alterations to progressive stages of cancer, and ILK has the potential as a serum biomarker for early-stage screening and for monitoring clinical status of patients after chemotherapy treatment. Inhibition of ILK activity in tumor cells reverses many of the malignant properties of tumor cells and as such ILK is a promising cancer therapeutic target. Previous studies have shown that the research of ILK is valuable, but the antibody of ILK must be purchased overseas, and its price is high, then the objective of our study is to prepare polyclonal anti-ILK antibody, With the antibody, ILK expression can be monitored throughout in vitro and in vivo gene transfer studies, the polyclonal antibody can be used in the further reseach with ILK gene.In the present study, first, we constructed the recombinant prokaryotig plasmid DNA expression vector encoding integrin-linked kinase (ILK). Fusion protein kd-ILK was expressed after IPTG induction and isolated and purified from the total proteins of E coli BL21 transformed by the recombinant plasmid pET32—ILK (1 — 1432); Then, the polyclonal antibody was got by immunizing rabbits with this purified fusion protein, and the quality of the antibody was identified. Second, we prepared the anti-polypeptide of ILK antibody, and purified...