A Novel Biochip System Focusing on Protein Detection by the Use of Labeled Glycopeptides Arrayed on a Novel Chip-material

摘要

The major goal of biochips is their use for rapid and economical characterization of biological samples. We have devoted considerable effort to the development of practical bio-detection systems. Most of protein-chips, in which proteins (antibodies) are immobilized as capture molecules, are impractical especially with regard to their stability. We have proposed a novel biochip-concept for a protein detection system involving labeled structured peptides as capture molecules that are arrayed on the chip surface. Visualization of interactions is not in a "one to one" manner, but as bar-code like patterns with fluorescent intensities generating "protein fingerprints", while the protein-protein interaction can be mimicked by a protein-peptide interaction [reviewed in 1]. Peptide array has greater potential, as even after drying immobilized peptides recognized analytes in a dose dependent manner [2]. Important factors for peptide micro-arrays are the construction of peptide libraries, the use of chip materials with a suitable surface chemistry, the deposition of peptide solutions by an arrayer, detection and data mining. In practice our system has been shown to discriminate the structures of a number of different proteins. As several toxic proteins, such as Ricinus communis agglutinin (RCA), cholera toxin, staphylococcal enterotoxin B, and Pseudomonas aeruginosa lectin (PA-I) recognize cell-surface carbohydrates of the host cells, these carbohydrate-binding species with protein-structure recognition may be applied for detection. The present paper describes the construction of glycopeptide libraries in addition to fluorescently labeled structured peptides (α-helical, P-loop and p-strand) and the development of the novel chip-material made from amorphous carbon with a special surface chemistry. The loading amounts on this amino-carbon plate were calculated ca 4 pmol/mm~2 [3]. This novel microarray, designated PepTenChip~R, satisfies all the requirements for protein detection, specificity, reproducibility, sensitivity, easy handling, stability (storage/transport) and production economics.