Functionalized Graphene Oxide as a Nanocarrier in a Multienzyme Labeling Amplification Strategy for Ultrasensitive Electrochemical Immunoassay of Phosphorylated p53 (S392)

Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, College of Chemistry, Central China Normal University, Wuhan, People's Republic of China.
Analytical Chemistry (Impact Factor: 5.83). 02/2011; 83(3):746-52. DOI: 10.1021/ac101715s
Source: PubMed

ABSTRACT P53 phosphorylation plays an important role in many biological processes and might be used as a potential biomarker in clinical diagnoses. We report a new electrochemical immunosensor for ultrasensitive detection of phosphorylated p53 at Ser392 (phospho-p53(392)) based on graphene oxide (GO) as a nanocarrier in a multienzyme amplification strategy. Greatly enhanced sensitivity was achieved by using the bioconjugates featuring horseradish peroxidase (HRP) and p53(392) signal antibody (p53(392)Ab(2)) linked to functionalized GO (HRP-p53(392)Ab(2)-GO) at a high ratio of HRP/p53(392)Ab(2). After a sandwich immunoreaction, the HRP-p53(392)Ab(2)-GO captured onto the electrode surface produced an amplified electrocatalytic response by the reduction of enzymatically oxidized thionine in the presence of hydrogen peroxide. The increase of response current was proportional to the phospho-p53(392) concentration in the range of 0.02-2 nM with the detection limit of 0.01 nM, which was 10-fold lower than that of the traditional sandwich electrochemical measurement for p53(392). The amplified immunoassay developed in this work shows acceptable stability and reproducibility, and the assay results for phospho-p53(392) spiked in human plasma also show good recovery (92-103.8%). This simple and low-cost immunosensor shows great promise for detection of other phosphorylated proteins and clinical applications.

  • Fullerenes Nanotubes and Carbon Nanostructures 01/2014; 23(5):410-417. DOI:10.1080/1536383X.2014.924509 · 0.64 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: This study reports on a novel biosensor for K-ras mutation gene detection by using hairpin locked nucleic acids (LNA) probes dually labeled with amines groups and biotin. The hairpin LNA probes was covalently immobilized on poly-Eriochrome cyanine R (ECR) film modified glassy carbon electrode (GCE) as a switch. Streptavidin-horseradish Peroxidase labeled gold nanoparticles (SA-HRP-AuNPs) bioconjugate were used as tracer for signal amplification. The immobilized hairpin LNA probe was in the "closed" state in the absence of K-ras mutation gene, which shielded biotin from being approached by the bulky SA-HRP-AuNPs bioconjugate due to the steric effect. When hairpin LNA probes hybridized with K-ras mutation gene, hairpin LNA probes undergo significant conformational change, forcing biotin away from GCE surface. As a result, the biotin label becomes accessible by the SA-HRP-AuNPs bioconjugate, and K-ras mutation gene hybridization event can be sensitively transduced via the numerous HRP enzymatically amplified electrochemical current signal. This new biosensor had a good specificity to distinguish the K-ras mutations, K-ras wild-type sequence, and a high sensitivity.
    Electrochimica Acta 10/2013; 108:808-813. DOI:10.1016/j.electacta.2013.07.042 · 4.09 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Here a new electrochemical immunoassay platform has been developed for the trace amounts of myelocytomatosis (c-Myc) oncoprotein detection to solve the urgent need for improved early diagnostic tools of cancer. To our knowledge, this is the first electrochemical immunosensor for c-Myc oncoprotein determination by employing gold nanoparticles as labels in a signal enhancement strategy. It is based on a sandwich immunoassay where the target c-Myc oncoprotein (CAg) is captured by the primary c-Myc antibody (CAb) modified on gold substrate, followed by adding another CAb conjugated to gold nanoparticle tags. The proposed sensor shows a linear range between 4.3 pmol L−1 and 43 nmol L−1, with an estimated detection limit of 1.5 pmol L−1. The recovery was achieved between 96.2% and 109% in 1% serum samples. This sensitive immunosensor holds great promise for the early diagnostic application of cancer at a curable stage.
    Sensors and Actuators B Chemical 05/2013; 181:835-841. DOI:10.1016/j.snb.2013.02.063 · 3.84 Impact Factor


Available from
May 20, 2014