SOLID PHASE GLYCAN AND GLYCOPEPTIDE ANALYSIS AND MICROFLUIDIC CHIP FOR GLYCOMIC EXTRACTION, ANALYSIS AND METHODS FOR USING SAME

摘要

Highly specific and novel solid phase methods for analyzing glycans and proteoglycans using a solid phase system are provided. The present invention also provides an integrated apparatus and methods of use which comprises a high-throughput glycan isolation and reverse-phase liquid chromatography (RPLC) for on-chip glycan extraction, modification and separation. The coverage of detected N-glycans by the GIG-chip-LC apparatus of the present invention can be significantly improved, especially for the low abundant species. Chip-LC by PGC minimizes dynamic range of glycan concentrations in fractions, resulting in detection of low-abundance glycans. Glycan isomers were able to be separated by the chip-LC portion of the apparatus. The GIG-chip-LC apparatus of the present invention can be used to analyze glycans from tissue and sera samples, thus providing a reliable tool for glycomic analysis. The reproducible performance and ability to detect unique glycans from tissue samples provides a powerful means for discovery of abnormal glycans associated with disease states.

主权项

1. A method of isolating glycans from glycoproteins in a biological sample comprising:
a) obtaining a biological sample comprising glycoproteins; b) denaturing the sample of a) to denature the glycoproteins and/or glycopeptides; c) conjugating the denatured glycoproteins and/or glycopeptides from b) with reductive amination to aldehyde groups on solid support; d) blocking unreacted aldehyde groups on the solid support with reductive amination; e) removing unconjugated glycoproteins and/or glycopeptides and remaining components; f) releasing the glycans from the glycoproteins and/or glycopeptides bound to the solid support of c); and g) isolating the glycans released from f).