Research Article Órpez-Zafra, Pavía, Pinto-Medel et al.
400 p = 0.000000197
300
200
100
0 Healthy controls Non-treated MS
Figure 3. ELISA determination of sIFNAR2 concentration in serum of non-treated MS patients and healthy controls.
ity of the assay for analyzing archival blood samples up to this time. Once the protocol was validated, we measured sIF-
NAR2 in serum from MS patients and HC. An abun- dance of soluble circulating cytokine receptors has been reported, with sIFNAR2, among them [14,26]. Almost all samples analyzed had detectable sIFNAR2 levels within the range of our ELISA, explained by the fact that the IFN-β receptor (IFNAR) is ubiquitously expressed on most cell types [4,27] and sIFNAR2 is an isoform resulting from alternative processing of the human IFNAR2 gene product [28,29]. The sIFNAR2 measurements with our validated
ELISA showed that MS patients, who had not been treated with IFN-β had lower circulating sIFNAR2 levels than HC. While developing the validation of the assay, a commercial kit for sIFNAR2 was released. Our recombinant sIFNAR2 was detected in the com- mercially available sIFNAR2 ELISA kit, and the standard protein of this commercial kit was detect- able with our ELISA. Moreover, some of the samples were analyzed with both ELISAs, showing that differ- ences between MS patients and HC followed the same pattern. To our knowledge, only one previous study found increased sIFNAR2 levels in IFN-β-treated patients compared with HC by ELISA [30], although details of the method and the standards used were not described.
2878 Bioanalysis (2015) 7(22) The present study demonstrated a significant
increase in serum sIFNAR2 in HC compared with untreated MS patients, with a slight degree of overlap- ping in the distribution of the marker in both popu- lations. Additional studies are needed to evaluate the role of sIFNAR2 as a useful biomarker for the diag- nosis or differential diagnosis in MS. If so, it will have the advantage of being detected in serum, unlike most accurate diagnostic markers previously described in MS such as oligoclonal IgG bands [31,32], free kappa light chain [32] or chitinase 3-like 1 [33], which are found in cerebrospinal fluid.
Conclusion The validation data characterize this ELISA as a suit- able method for the quantification of the soluble iso- form of the IFNAR2 receptor in human serum. The availability of this sandwich ELISA will provide con- sistent results for assessment of sIFNAR2 concentra- tions in MS, representing a practical tool for further studies aimed to explore the implication of sIFNAR2 in MS and other immunological diseases.
Future perspective Soluble receptors of cytokines normally participate in the control of cytokine activity and the ability of bind- ing to the cytokine have prompted interest in their use as therapeutic agents. However, the soluble isoform of
future science group
sIFNAR2 (ng/ml)
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