The monoclonal antibody 26 raised against tetanus toxoid also recognizes tetanus toxin and β2-glycoprotein I - its binding properties in vitro and potential applications

• Published in: Journal of the Serbian Chemical Society 2009, Vol.74 (3), p.245-257
• Main Authors: Inic-Kanada, Aleksandra, Stojanovic, Marijana, Zivkovic, Irena, Petrusic, Vladimir, Dimitrijevic, Ljiljana
• Format: Article
• Language: English
• Subjects: formaldehyde; monoclonal antibodies; tetanus toxin; tetanus toxoid
• ISSN: 0352-5139; 1820-7421
• DOI: 10.2298/JSC0903245I

A murine monoclonal IgG1 antibody, marked as MAb26, specific for tetanus toxoid has been immunochemically characterized. By performing enzyme-linked immunosorbent assays (ELISAs) and western blot analyses, it was demonstrated that MAb26 reacted with tetanus toxoid, tetanus toxin and ?2-glycoprotein I (?2GPI). According to the results, MAb26 recognized the sequential epitope on the tetanus heavy chain. The affinity constant, calculated from Scatchard plots of MAb26 binding to tetanus toxoid, was 1.145?108 M-1 and the measurement of the relative affinity of MAb26 by ELISA using thiocyanate elution showed a significantly higher affinity of MAb26 to the toxoid (p = 0.0012) in comparison to the toxin. Additionally, the reactivity of MAb26 toward the toxoid forms increased when the tetanus toxin was detoxified using 8 mM and higher formaldehyde concentrations. The similarity of the tetanus toxoid to several sera proteins, either at the level of its conformation (IL-1?) or at the level of peptide sequences (?2GPI, laminin) favors its role in autoimmunity by the mechanism of molecular mimicry. As the induction of an autoimmune disease is dependent on the breakdown of tolerance, which could be the result of an overt hyperstimulation, the control of the presence and concentration of self-reactive epitopes in vaccine preparations is a prerequisite. In this study, it was shown that MAb26 can: 1) discriminate between the tetanus toxin and different toxoid forms, which makes it a good candidate for antibody control during vaccine preparation; 2) due to its cross-reactivity with ?2GPI, it could provide information on the presence of a potentially dangerous sequential epitope expressed at the protein surface. Ovaj rad opisuje imunohemijsku karakterizaciju misjeg IgG1 monoklonskog antitela oznacenog kao MAt26. Enzimskim imunosorbentnim testom (ELISA) i Western blot analizom je pokazano da MAt26 reaguje sa tetanus toksoidom, tetanus toksinom i ?2-glikoproteinom I (?2GPI). Prema nasim rezultatima, MAt26 prepoznaje sekvencioni epitop na teskom lancu molekula tetanusa. Konstanta afiniteta MAt26 za tetanus toksoid, izracunata na osnovu Skacardovog dijagrama, je 1,145?108 M-1. Na osnovu elucije tiocijanatom, koriscene za odredjivanje relativnog afiniteta MAt26 za tetanus toksin i tetanus toksoid, postupkom baziranim na ELISA-i, pokazan je znatno veci (p = 0,0012) afinitet MAt26 ka toksoidnoj formi. Takodje, reaktivnost MAt26 ka toksoidnoj formi rasla je sa porastom koncentracije