title: Semi-quantitative measurement of a specific glycoform using a DNA-tagged antibody and lectin affinity chromatography for glyco-biomarker development.
abstract: Aberrant glycosylation-targeted disease biomarker development is based on cumulative evidence that certain glycoforms are mass-produced in a disease-specific manner. However, the development process has been hampered by the absence of an efficient validation method based on a sensitive and multiplexed platform. In particular, ELISA-based analytical tools are not adequate for this purpose, mainly because of the presence of a pair of N-glycans of IgG-type antibodies. To overcome the associated hurdles in this study, antibodies were tagged with oligonucleotides with T7 promoter and then allowed to form a complex with corresponding antigens. An antibody-bound specific glycoform was isolated by lectin chromatography and quantitatively measured on a DNA microarray chip following production of fluorescent RNA by T7-trascription. This tool ensured measurement of targeted glycoforms of multiple biomarkers with high sensitivity and multiplexity. This analytical method was applied to an in vitro diagnostic multivariate index assay where a panel of hepatocellular carcinoma (HCC) biomarkers comprising alpha-fetoprotein, hemopexin, and alpha-2-macroglobulin (A2M) was examined in terms of the serum level and their fuco-fractions. The results indicated that the tests using the multiplexed fuco-biomarkers provided improved discriminatory power between non- hepatocellular carcinoma and hepatocellular carcinoma subjects compared with the alpha-fetoprotein level or fuco-alpha-fetoprotein test alone. The developed method is expected to facilitate the validation of disease-specific glycan biomarker candidates.
authors.affiliation: From the ‡Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon 305-806, Korea; §Korea University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon 305-350, Korea;
authors.forename: Ju Hee
authors.initials: JH
authors.lastname: Lee
authors.affiliation: From the ‡Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon 305-806, Korea;
authors.forename: Chang Hee
authors.initials: CH
authors.lastname: Cho
authors.affiliation: From the ‡Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon 305-806, Korea;
authors.forename: Sun Hee
authors.initials: SH
authors.lastname: Kim
authors.affiliation: From the ‡Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon 305-806, Korea;
authors.forename: Jeong Gu
authors.initials: JG
authors.lastname: Kang
authors.affiliation: ¶Division of Mass Spectrometry, Korea Basic Science Institute, Ochang-Myun, Cheongwon-Gun 363-883, Korea; ‖GRAST, Chungnam National University, Daejeon 305-764, Korea;
authors.forename: Jong Shin
authors.initials: JS
authors.lastname: Yoo
authors.affiliation: **Department of Laboratory Medicine, School of Medicine, Pusan National University, Busan 609-735, Korea.
authors.forename: Chulhun Ludgerus
authors.initials: CL
authors.lastname: Chang
authors.affiliation: From the ‡Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon 305-806, Korea; §Korea University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon 305-350, Korea; omsys1@kribb.re.kr jhko@kribb.re.kr.
authors.forename: Jeong-Heon
authors.initials: JH
authors.lastname: Ko
authors.affiliation: From the ‡Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon 305-806, Korea; §Korea University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon 305-350, Korea; omsys1@kribb.re.kr jhko@kribb.re.kr.
authors.forename: Yong-Sam
authors.initials: YS
authors.lastname: Kim
mesh_terms: D000906:Antibodies / Q000737:chemistry* / Q000276:immunology; D014408:Biomarkers, Tumor / Q000737:chemistry / Q000302:isolation & purification*; D006528:Carcinoma, Hepatocellular / Q000097:blood / Q000175:diagnosis*; D045744:Cell Line, Tumor; D002846:Chromatography, Affinity / Q000379:methods; D004247:DNA / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification*; D006801:Humans; D037102:Lectins / Q000737:chemistry; D008113:Liver Neoplasms / Q000097:blood / Q000175:diagnosis*; D020411:Oligonucleotide Array Sequence Analysis; D057089:Pathology, Molecular / Q000379:methods
chemical_list: D000906:Antibodies; D014408:Biomarkers, Tumor; D006023:Glycoproteins; D037102:Lectins; D004247:DNA