| Project/Area Number |
09557220
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
Laboratory medicine
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| Research Institution | Osaka Medical College |
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Principal Investigator |
SHIMIZU Akira Faculty of Medicine, Professor, 医学部, 教授 (00028581)
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| Co-Investigator(Kenkyū-buntansha) |
YAGI Masayuki Arkray Co., Ltd., Director, 技術研究部, 係長
KISHIKAWA Masahiko Faculty of Medicine, Osaka Medical Research Assistant, 医学部, 助手 (40268199)
NAKANISHI Toyofumi Faculty of Medicine, Osaka Medical College Associate Professor, 医学部, 講師 (10247843)
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| Project Period (FY) |
1997 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥11,600,000 (Direct Cost: ¥11,600,000)
Fiscal Year 2000: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 1999: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 1998: ¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 1997: ¥5,000,000 (Direct Cost: ¥5,000,000)
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| Keywords | HbAlc / electrospray ionization / mass spectrometry / abnormal Hb / HPLC / immunoassays / Glu-C / ESI / MS / 蛋白分解酵素 / デイテクターゲイン / 固相化酵素 / 糖化ヘモグロビン / HbAlc / 液体クロマトグラフィー / 異常Hb / V8プロテアーゼ / ホルミルHb / カルボキシ末端アルギニン欠損Hb / HPLC / 方法間誤差 / ESI / MS / β鎖糖化成分 / カルバミルHb |
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| Research Abstract |
We established a reference method for glyco-haemoglobin (HbAlc), which is a most popular index for control of diabetes mellitus. The N-terminal residues of the Hb β-chains, which are released by enzymatic cleavage of the intact Hb molecule with endoproteinase Glu-C, were measured by electrospray ionization mass spectrometry(ESI/MS). The procedure was originally described by Kobolt et al., and we modified several points. We calculated the peak ratio of glycated to non-glycated hexapeptide by monitoring both singly and doubly protonated ions ('weighed sum method'). Secondly, We prepared a calibrator by mixing synthetic peptides, instead of mixing purified HbAlc and HbA_0. The coefficient of variation (CV) of intra- and inter-assay by our method was less than 2.0%. We studied the structure of various abnormal Hbs also using ESI/MS Most cases were requested to elucidate the cause of the unexpected values of HbAlc measured by HPLC.We measured HbAlc in samples with abnormal Hbs using 3 commercial systems and the ESI/MS method, and we clarified the error of HbAlc values both by HPLC and immunoassays for 25 kinds, 49 cases with abnormal Hbs.
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