Combining peptide and DNA for protein assay: CRIP1 detection for breast cancer staging.

In this work, a novel method for a protein assay is proposed which uses the specific protein-binding peptide of the target protein and sequence-specific DNA to interact with the target as the capture and detection probe, respectively. Meanwhile, since the DNA sequence can be coupled with gold nanoparticles to amplify the signal readout, a sensitive and easily operated method for protein assay is developed. We have also employed a transcription factor named as cysteine-rich intestinal protein 1 (CRIP1), which has been identified as an ideal biomarker for staging of breast cancer, as the model protein for this study. With the proposed method, CRIP1 can be determined in a linear range from 1.25 to 10.13 ng/mL, with a detection limit of 1.25 ng/mL. Furthermore, the proposed method can be directly used to assay CRIP1 in tissue samples. Owing to its desirable sensitivity, excellent reproducibility, and high selectivity, the proposed method may hold great potential in clinical practice in the future.

Highly sensitive protein detection based on a novel probe with catalytic activity combined with a signal amplification strategy: assay of MDM2 for cancer staging.

A novel probe with catalytic activity is used for highly sensitive MDM2 detection.

Design of a bi-functional peptide for protein assays: observation of cortactin expression in human placenta.

A bi-functional peptide is designed to incorporate protein recognition and signal amplification functions into a single short peptide sequence.

A novel method to assay molecular chaperone activity of HSP70: evaluation of drug resistance in cancer treatment.

In this work, we have proposed a novel method to specifically assay the molecular chaperone activity of HSP70 based on the HSP70-substrate peptide interaction. By selectively labeling the substrate peptide of HSP70 via host-guest interaction with two different cucurbituril species, the HSP70-substrate peptide interaction can be transduced into detectable signal readout. By using the signal readout, assay of the molecular chaperone activity of HSP70 can be achieved. Moreover, by using our method, chaperone activity of HSP70 can serve as a reliable indicator of drug resistance in cancer treatment. The experimental results reveal that enhanced chaperone activity of HSP70 is observed in both drug-resistant cancer cell line and the serum of cancer victim subject to anti-cancer therapy. Therefore, the proposed method to assay the molecular chaperone activity of HSP70 can be a tool of efficiency in evaluating therapeutic response in HSP70-targeted cancer treatment.

A general way to assay protein by coupling peptide with signal reporter via supermolecule formation.

Protein-binding peptide is recently recognized as an effective artificial affinity reagent for protein assays. However, its application is hampered by the limited choices of available signal readout methods. Herein, we report a general electrochemical signal readout method for protein-binding peptides exploiting the host-guest chemistry of cucurbituril. Via the formation of supermolecules among cucurbituril, electrochemical reporter, and the peptide, a protein-binding peptide can be noncovalently coupled with the electrochemical reporter. To assay the target protein, the protein-binding peptides are first self-assembled in the sensing layer, and after the capturing of the target protein, a portion of the peptides become protein-bound. The protein-free peptides are then coupled with the electrochemical reporter to yield a signal readout inversely proportional to the amount of the captured target proteins. Since the only requirement of supermolecule formation is the incorporation of aromatic amino acids in the peptide sequence, this strategy is universally applicable to many protein-binding peptides. The generality and target specificity of the proposed method are successfully demonstrated in the assays of two kinds of target proteins: tumor necrosis factor-α and amyloid ß 1-42 soluble oligomer, respectively. The feasibility of our method is also tested in the monitoring of tumor necrosis factor-α secretion activity of HL-60 cells. These results indicate that our method can have great use in protein detection in the future.