title: Mass Spectrometric N-Glycan Analysis of Haptoglobin from Patient Serum Samples Using a 96-Well Plate Format.
abstract: Alterations in glycosylation of serum glycoproteins can provide unique and highly specific fingerprints of malignancy. Our previous mass spectrometric study revealed that the bifucosylation level of serum haptoglobin was distinctly increased in hepatocellular carcinoma (HCC) patients versus liver cirrhosis of all three major etiologies. We have thus developed a method for the analysis of large numbers of serum samples based on a 96-well plate platform for the evaluation of fucosylation changes of serum haptoglobin between HCC versus cirrhosis. Haptoglobin was isolated from the serum of individual patient samples based on an HPLC column immobilized with antihaptoglobin antibody via hydrazide immobilization chemistry. Only 10 μL of serum was required for glycan extraction and processing for MALDI-QIT mass spectrometry analysis using the 96-well plate format. The bifucosylation degrees of haptoglobin in individuals were calculated using a quantitative glycomics method. The MS data confirmed that the bifucosylated tetra-anntenary glycan was upregulated in HCC samples of all etiologies. This study provides a parallel method for processing glycan content for haptoglobin and evaluating detailed changes in glycan structures for a potentially large cohort of clinical serum samples.
authors.affiliation: Department of Surgery, University of Michigan Medical Center , 1150 West Medical Center Drive, Ann Arbor, Michigan 48109, United States.
authors.forename: Jianhui
authors.initials: J
authors.lastname: Zhu
authors.affiliation: Department of Surgery, University of Michigan Medical Center , 1150 West Medical Center Drive, Ann Arbor, Michigan 48109, United States.
authors.forename: Jing
authors.initials: J
authors.lastname: Wu
authors.affiliation: Department of Surgery, University of Michigan Medical Center , 1150 West Medical Center Drive, Ann Arbor, Michigan 48109, United States.
authors.forename: Haidi
authors.initials: H
authors.lastname: Yin
authors.affiliation: Department of Internal Medicine, UT Southwestern Medical Center , 5323 Harry Hines Boulevard, Dallas, Texas 75390, United States.
authors.forename: Jorge
authors.initials: J
authors.lastname: Marrero
authors.affiliation: Department of Surgery, University of Michigan Medical Center , 1150 West Medical Center Drive, Ann Arbor, Michigan 48109, United States.
authors.forename: David M
authors.initials: DM
authors.lastname: Lubman
mesh_terms: D000368:Aged; D000437:Alcoholism / Q000097:blood* / Q000150:complications / Q000235:genetics / Q000473:pathology; D015415:Biomarkers / Q000097:blood; D002240:Carbohydrate Sequence; D006528:Carcinoma, Hepatocellular / Q000097:blood* / Q000209:etiology / Q000235:genetics / Q000473:pathology; D002851:Chromatography, High Pressure Liquid; D005260:Female; D005643:Fucose / Q000378:metabolism; D006031:Glycosylation; D006242:Haptoglobins / Q000235:genetics / Q000378:metabolism; D006509:Hepatitis B / Q000097:blood* / Q000150:complications / Q000235:genetics / Q000473:pathology; D006526:Hepatitis C / Q000097:blood* / Q000150:complications / Q000235:genetics / Q000473:pathology; D006801:Humans; D008103:Liver Cirrhosis / Q000097:blood* / Q000150:complications / Q000235:genetics / Q000473:pathology; D008113:Liver Neoplasms / Q000097:blood* / Q000209:etiology / Q000235:genetics / Q000473:pathology; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D009367:Neoplasm Staging; D010449:Peptide Mapping; D011499:Protein Processing, Post-Translational; D059748:Proteolysis; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D014357:Trypsin / Q000737:chemistry
chemical_list: D015415:Biomarkers; D006242:Haptoglobins; D005643:Fucose; D014357:Trypsin
keywords: 96-well plate format; HPLC; MALDI; glycan; haptoglobin; hepatocellular carcinoma