Sensitive chemiluminescence detection of wild-type p53 protein captured by surface-confined consensus DNA duplexes.

A novel chemiluminescence (CL) biosensor for sensitive detection of wild-type p53 protein has been proposed. The wild-type p53 protein in solution was captured by highly specific consensus double-stranded (ds) oligonucleotides (ODNs) preimmobilized onto a gold plate. The cysteine residues on the exterior of the wild-type p53 molecules were then derivatized with N-biotinoyl-N'-(6-maleimidohexanoyl) hydrazide (biotin-Mi) for the attachment of streptavidin-horseradish peroxidase (SA-HRP) complex. The attached HRP molecules could catalyze the CL reaction between luminol and H2O2, producing an enhanced CL signal. The CL intensity was dependent on the surface coverage of the HRP molecules, which was related to the concentration of wild-type p53 protein. Under the optimal experimental conditions, the CL intensity increased linearly with the concentration of wild-type p53 protein from 0.01 to 0.5nM. The detection limit was estimated to be 3.8pM. The proposed method has been successfully utilized for the assay of wild-type p53 protein in normal and cancer cell lysates. The sensing protocol is sensitive, cost-effective, and holds great promise for clinical diagnosis.