Immune recognition of recombinant parvovirus B19 non-structural (rNS1) protein was studied by immunoblot and lymphoproliferative assays in blood from the following B19 seropositive groups: B19 infected (n = 14), B19 exposed but non-infected (n = 16), other illness with rash (n = 3), chronic arthropathy of unknown aetiology (n = 4) and healthy controls (n = 7). Sera from 11 B19 seronegative subjects were also studied. Sera collected at initial diagnosis or at the time of accidental B19 exposure in pregnancy were tested for NS1 antibody and evidence of B19 DNA by nested PCR. Follow-up specimens were obtained 3-12 months later for serological, PCR and proliferation studies. B19 DNA was detected sporadically in early specimens and in one follow-up specimen from a subject who developed chronic arthropathy after B19 infection. There was no correlation with development of arthropathy. NS1-specific IgG was detected in early sera from B19-infected and exposed subjects but to a lesser degree in follow-up specimens, and in only one healthy control serum. No correlation with the presence of NS1-specific antibodies was found with development of acute or chronic arthropathy. Although lymphocyte proliferation in response to stimulation with rNS1 in vitro occurred at a higher frequency in patients who developed acute and chronic joint manifestations after B19 infection, suggesting an association with this outcome, NS1-reactive lymphocytes were also found in three B19 seronegative patients, two of whom had recently been exposed to B19 but had no illness. Hence, immune recognition of NS1 may be more indicative of recent infection with, or exposure to, parvovirus B19 than associated with development of arthropathy as previously reported.