Anti-Interferon beta (IFNbeta) antibodies are frequently produced during treatment with IFNbeta in Multiple Sclerosis (MS) patients. In recent years, it has become clear that these antibodies cause a decrease in IFNbeta-induced biomarkers and in IFNbeta clinical efficacy. Anti-IFNbeta antibodies are mainly of the IgG isotype, which consists of 4 subclasses. In this study, we tested whether changes occurred in IgG subclasses over time. A series of sera from 21 IFNbeta-treated patients (11 IFNbeta-1a, 10 IFNbeta-1b) were analysed longitudinally using a capture ELISA. IFNbeta-1a treated patients had a restricted subclass distribution, whilst IFNbeta-1b-treated patients demonstrated a wider distribution. When compared to IFNbeta-1b-treated patients, IFNbeta-1a-treated patients had lower levels of total and subclass-specific IgGs against IFNbeta. In particular, antibody levels were markedly lower in the neutralizing antibody (NAb) negative (-) category of IFNbeta-1a-treated patients in comparison to the NAb-IFNbeta-1b-treated patients. The most striking observation of this study were the very low levels or complete absence of IgG3 subclass-specific antibodies to IFNbeta in IFNbeta-1a-treated patients. This difference in the levels of IgG3 may help to clarify the differences in the overall pattern of development of anti-IFNbeta antibodies in IFNbeta-1a-and IFNbeta-1b-treated patients.