Figure 1.
Specificity of the polyclonal antibody against native hepcidin.
(A) Immunohistochemical staining of liver tissue sections using the polyclonal antibody against hepcidin25-His (a-Hep25). Secondary anti-rabbit antibody was used as negative control (negative). The specificity of the polyclonal antibody was verified after reduction of the signal following preincubation with hepcidin25-His. (B) Western blot analysis of serum proteins less than 30 kDa. 10 ml of serum was filtered through a 30 kDa filter, and the filtrate was precipitated with 25% TCA. Precipitated proteins were subjected to electrophoresis on a 4–12% Nu-PAGE gel, followed by Western blot using the polyclonal antibody against hepcidin25-His.
Figure 2.
Representative calibration curve for recombinant hepcidin25-His.
The range of the assay is 10–1500 µg/L.
Table 1.
Intra-assay variation.
Table 2.
Inter-assay variation.
Table 3.
Recovery of calibrator added to human serum samples.
Table 4.
Dilution linearity of ELISA.
Figure 3.
Hepcidin serum concentration in healthy controls (control), patients with iron deficiency anemia (IDA), juvenile haemochromatosis (JH) and Hodgkin's lymphoma (HL).
Box plots show the 25th and 75th percentile with median value for each group. Minimum and maximum values are also depicted. Significant difference compared to control is indicated by asterisk (**p<0.010).
Figure 4.
Correlation between serum hepcidin and ferritin in healthy controls.
Hepcidin values measured by our ELISA assay correlate significantly with ferritin levels. Pearson correlation: 0.474 (p = 0.006).
Figure 5.
Correlation between serum hepcidin measured by our ELISA assay and by the SELDI-TOF-MS method.
Hepcidin values of 6 serum samples, measured by the two methods correlate significantly. Pearson correlation: 0.863 (p = 0.027).