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prca1481-sup-0001-Tables.doc90K

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

prca1481-sup-0002-Figures.ppt965K

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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