Establishment of the biotin-streptavidin ELISA for detection of serum connective tissue growth factor and its preliminary clinical application
10.3760/cma.j.issn.1009-9158.2011.11.008
- VernacularTitle:生物素-链霉亲和素ELISA检测血清CTGF方法的建立及其初步应用
- Author:
Dai ZHANG
;
Nianyue WANG
;
Weihong REN
;
Wei ZHAO
;
Songren ZHAO
- Publication Type:Journal Article
- Keywords:
Connective tissue growth factor;
Liver cirrhosis;
Hepatitis B,chronic;
Enzymelinked immunosorbent assay
- From:
Chinese Journal of Laboratory Medicine
2011;34(11):993-998
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a biotin-streptavidin ELISA method to measure CTGF,and evaluate the clinical value of CTGF for the diagnosis of liver fibrosis in chronic hepatitis B (CHB) patients.Methods Biotinylated anti-CTGF polyclonal antibody and monoclonal antibodies were prepared for the establishment of this biotin-streptavidin ELISA method.Two hundreds and sixty-four CHB patients were subjected into non-or mild liver fibrosis group (S0-S1,108 cases) and severe liver fibrosis group (S2-S4,156 cases),according to the liver biopsy pathological diagnosis.The CTGF assay's diagnostic capacity for CHB was assessed by comparing the area under ROC (AUC) with that of a panel of hepatic fibrosis markers ( HA,PC Ⅲ,C Ⅳ,LN and APRI).Analysis of variance and rank sum test were performed to carry out comparisons between multiple groups.Student's t-test and Mann-Whitney U test were performed for the pairwise comparison between multiple samples.Spearman rank correlation test was performed to analyze the correlation between different hepatic fibrosis stages.Results The minimum detectable dose and detection rang of the ELISA was 0.2 μg/L and 0-64 μg/L respectively.The intra-assay and inter-assay CV at high and low level were 4.5％,9.8％ and 10.1％,12.8％ respectively.Serum CTGF concentrations in S0-S1 group and S2-S4 group were 6.7(3.1 - 10.1 ) μg/L and 16.1 ( 11.8 -27.2) μg/L,with a statistically significant difference (U =1 217,P ＜0.001 ).There was a significant correlation between the levels of serum CTGF and fibrosis stages ( r =0.689,P ＜ 0.001 ),AUC of CTGF was 0.841 (95％ CI:0.762 - 0.920) in distinguishing mild fibrosis from significant fibrosis.When the cut-off value of CTGF was 10.3 μg/L,the sensitivity and specification was 70.5％ and 82.4％ respectively.The sensitivity of parallel combination test of CTGF and APRI was 96.1％,which was higher than that of HA (75.6％),PC Ⅲ (70.5％ ),C Ⅳ(63.6％),LN(79.5％ ),APRI(86.3％ ).The specificity of the combination test was 65.5％,which was lower than of above liver fibrosis markers [HA ( 72.5％ ),PC Ⅲ ( 76.5％ ),C Ⅳ ( 78.4％ )].The specificity of serial combination test of CTGF and PC Ⅲ was 95.9％,which was higher than that of HA,PC Ⅲ,CⅣ,LN(64.7％ ),APRI(66.1％ ),however,the sensitivity of the combination test was 67.7％,which was lower than that of above HA,PC Ⅲ,and APRI.Conclusions The biotin-streptavidin ELISA method measuring serum CTGF has a high minimu detectable dose sensitivity,and specificity.Serum CTGF level is significantly correlated with fibrosis stage,and CTGF maybe a valuable marker for liver fibrosis assessment.The paralledl combination of CTGF and APRI could be used as screening for significant liver fibrosis markers.The serial combination of CTGF and PC Ⅲ may be considered as a confirmatory diagnostic marker for liver fibrosis.
