Serum Mac-2 binding protein levels as a novel diagnostic biomarker for prediction of disease severity and nonalcoholic steatohepatitis
Version of Record online: 17 SEP 2013
© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
PROTEOMICS - Clinical Applications
Special Issue: Glycan Biomarker Discovery
Volume 7, Issue 9-10, pages 648–656, October 2013
How to Cite
Kamada, Y., Fujii, H., Fujii, H., Sawai, Y., Doi, Y., Uozumi, N., Mizutani, K., Akita, M., Sato, M., Kida, S., Kinoshita, N., Maruyama, N., Yakushijin, T., Miyazaki, M., Ezaki, H., Hiramatsu, N., Yoshida, Y., Kiso, S., Imai, Y., Kawada, N., Takehara, T. and Miyoshi, E. (2013), Serum Mac-2 binding protein levels as a novel diagnostic biomarker for prediction of disease severity and nonalcoholic steatohepatitis. Prot. Clin. Appl., 7: 648–656. doi: 10.1002/prca.201200137
- Issue online: 9 OCT 2013
- Version of Record online: 17 SEP 2013
- Accepted manuscript online: 14 JUN 2013 09:59AM EST
- Manuscript Accepted: 30 MAR 2013
- Manuscript Revised: 28 FEB 2013
- Manuscript Received: 25 DEC 2012
- Ministry of Education, Culture, Sports, Science, and Technology of Japan
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Table S1. Distribution of parameters according to Matteoni's classification in the NAFLD patients
Table S2. Histological Characteristics
Table S3. Correlation coefficients of relationships between serum Mac-2bp levels and various parameters
Table S4. Multiple logistic regression analysis of factors associated with stage 2–4 NAFLD compared to stage 0–1 NAFLD
Figure S1. Preparation of recombinant human Mac-2bp and mouse monoclonal antibodies against human Mac-2bp.
(A) The structures of recombinant human Mac-2bp and the epitope mapping of 2 kinds of human Mac-2bp monoclonal antibodies (67A1, 8A2). Full and partial length recombinant human Mac-2bps containing the FLAG tag at the C terminus were described. Yellow circles demonstrate N-glycan binding sites of Mac-2bp according to its amino acid sequence. The amino acid numbers were described at the top figure of hM2BP. M2BP; Mac-2bp, Xa; the cleavage site of the factor Xa protease, TMB; tetramethylbenzidine, HRP; horseradish peroxidase.
(B) Recombinant human Mac-2bp (containing the FLAG tag at the C terminus) was used for immunoblotting using 2 kinds of mouse monoclonal antibodies against human Mac-2bp (67A1, 8A2) and against FLAG tag (FLAG). Arrows indicate Mac-2bp (* Lower molecular weight band indicates truncated form of Mac-2bp).
Figure S2. Mouse monoclonal antibodies (67A1, 8A2) bound to human serum Mac-2bp.
Twenty μL of two NASH patient sera (the serum Mac-2bp levels measured by ELISA kit were 5.0 (lane 1) and 3.0 μg/mL (lane 2), respectively) were applied for immunoblotting analysis, using 2 kinds of mouse monoclonal antibodies [(A) 67A1, (B) 8A2] after immunoprecipitation. Arrows indicate Mac-2bp (* Lower molecular weight band indicates truncated form of Mac-2bp).
Figure S3. A standard curve of Mac-2bp ELISA.
Recombinant Mac2-bp produced by HEK293 cells was used as standard. The concentration of Mac-2bp was described as μg/mL. Detail procedure was described in Materials and Methods section.
Figure S4. Correlation between serum Mac-2bp levels and liver fibrosis stage in NAFLD patients
(A) Serum Mac-2bp levels in each stage of liver fibrosis in NAFLD patients.
(B) Comparison of serum Mac-2bp levels between non-mild fibrosis and advanced fibrosis in NAFLD patients.
(C) ROC curves for Mac-2bp and the M30 antigen for discrimination of advanced liver fibrosis in NAFLD patients.
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