We did not detect significant differences in serum IgM to LC (IgM-LC) nor in IgG reactivity to PC (IgG-PC) or LC (IgG-LC) between patients with MS and controls (figure 1B and not shown)

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We did not detect significant differences in serum IgM to LC (IgM-LC) nor in IgG reactivity to PC (IgG-PC) or LC (IgG-LC) between patients with MS and controls (figure 1B and not shown). Open in a separate window Figure Mupirocin 1 Serum IgM-PC and IgM-LC in patients with MS and controls(A) Serum IgM-PC controls. patients with SPMS, PPMS, and BENMS and controls. MS and control samples did not differ in serum levels of IgM antibodies reactive with LC, nor in IgG antibodies reactive with LC or PC. Conclusions Serum IgM-PC antibodies are elevated in patients with MS, particularly during the CIS and RRMS phases of the disease. Thus, serum IgM-PC is a candidate biomarker for early inflammatory stages of MS. Classification of evidence This study provides Class III evidence that serum antibodies to PC are elevated in patients with MS. The study is rated Class III because of the case control design and the risk of spectrum bias: antibody levels in patients with MS were compared with healthy controls. The high prevalence of immunoglobulin G (IgG) and immunoglobulin M (IgM) oligoclonal bands in the CSF has guided multiple studies focused on the identification of antibody targets in MS.1,2 Lipids are a major component of myelin, the main target of the autoimmune response in MS.3 New antigen microarray techniques have allowed the detection of lipid-reactive antibodies in the CSF of patients with MS.4,5 Indeed, IgG antibodies to sulfatide, ganglioside GM4, and galactocerebroside6,C9 worsen disease pathogenesis in MS experimental models.4 In addition, patients with MS harboring CSF IgM reactive with phosphatidylcholine (PC) develop a more severe disease course.6,7,10,C12 Moreover, serum IgG antibodies to lactosylceramide (LC) are associated with cerebral tissue damage in patients with MS.13 These data suggest that lipid-reactive antibodies may contribute to disease pathogenesis and constitute useful as biomarkers in MS. In this study, we evaluated the potential of lipid-reactive antibodies as biomarkers in MS. We detected increased levels of serum IgM antibodies to PC (IgM-PC) in patients with MS. In contrast, we did not detect differences in serum IgG antibodies to PC (IgG-PC), nor IgM or IgG to LC (IgM-LC and IgG-LC, respectively) between MS and controls. Thus, serum IgM-PC is a candidate biomarker for MS. Methods Study design We developed an ELISA technique to study IgG and IgM antibodies reactive with LC and PC in patients with MS. This Class III retrospective cohort control study included individuals from different cohorts (United States and Spain). The study included a narrow spectrum of persons with MS, non-MS myelin diseases (Non-MSMYDs) and nonmyelin neurologic diseases (Non-MYNDs) with a retrospective longitudinal follow-up, and without neurologic disease (controls). The MS patient and control samples analyzed are described in table 1. The samples were analyzed by ELISA by an investigator blinded to disease status; the diagnostic test results and disease status were determined by different team members. Clinical Mupirocin information was provided from a clinical database of the different patients with connected biobanks (blood and CSF samples). Table 1 Demographic and medical data from individuals with MS, Non-MSMYD, and Non-MYND and control group Open in a separate window Patients Samples were from the Comprehensive Longitudinal Investigation of Multiple Sclerosis (CLIMB, Boston, MA) and EPIC (San Francisco, CA), as MAP2K7 part of the SUMMIT consortium14,15; from your Servicio de Neurologia Hospital Virgen del Rocio (Sevilla, Spain); from your Servicio de Inmunologa del Hospital Clnico San Carlos Mupirocin (Madrid, Spain); and from your Servicio de Neurologa del Hospital Universitario Quirnsalud (Madrid, Spain). The classification and medical data of all samples analyzed are summarized in table 1. Peripheral blood samples from individuals with MS (362), individuals with Non-MSMYD (10), individuals with Non-MYND (11), and from 80 individuals without neurologic diseases (control group) were collected. Individuals were free of relapses at the time of blood sample collection. Sera samples were aliquoted and stored at ?80C until analyzed. ELISA assay To detect IgG and IgM antibodies reactive with LC (Matreya, State College, PA) and Personal computer (Sigma-Aldrich, St. Louis, MO), 96-well plates Mupirocin were 1st coated with LC diluted in dimethyl sulfoxide and Personal computer diluted in methanol at 10 g/mL. After washing 3 times with phosphate-buffered saline, the wells were treated with obstructing solution. Serum samples were diluted (1/50, 1/100, 1/200) in obstructing answer and pipetted into the wells because we acquired the higher signal and a lower background. IgG or IgM antibodies were recognized with anti-human IgG biotin and anti-human IgM biotin, respectively (Jackson ImmunoResearch, Western Grove, PA), followed by avidinChorse peroxidase (Sigma-Aldrich). Finally, we used TMB-one (Kementec, Taastrup, Denmark) as substrate. Plates were go through at 450 nm using an Infinite 200 PRO spectrophotometer (Tecan instrument, Zrich, Switzerland). We classified a serum as positive when the optic denseness.