Human herpesvirus 6 (HHV-6) is one of many infectious agents that have been implicated in the pathogenesis of multiple sclerosis (MS). Intrathecal immunoglobulin (Ig) production with oligoclonal CSF bands are hallmarks of both MS and infections of the CNS. In neuroinfections the intrathecal Ig production is directed largely against the respective agent, while MS patients mount an intrathecal Ig production against different pathogens. In this study a total of 77 serum/CSF pairs were first analyzed for an intrathecal immune response against HHV-6. We found that 21% of the MS patients, but none of the control donors showed an intrathecal immunoglobulin (Ig) response against HHV-6 (p = 0.017). Patients with such an intrathecal Ig production had a significantly higher total amount of Ig (p = 0.007) and a higher cell number in the CSF (p = 0.03). In a second step, four of the MS-patients who showed a strong intrathecal Ig production against HHV-6 were examined for immune reactivity against other viruses (human herpesvirus type 1, measles virus, human cytomegalovirus, rubella virus, varicella zoster virus). All of these patients had an intrathecal Ig production against at least one other, unrelated virus. Together, our findings strongly argue that in the majority of MS patients without an intrathecal Ig response to HHV-6 (about 80 % in our study), this virus is not involved in the pathogenesis. In the other 20% the intrathecal Ig production to HHV-6 might reflect reactivation of HHV-6 or could be part of a polyspecific B cell activation.