Epidemiological and virological evidence suggests that invasive squamous cell carcinoma (SCC) of the vulva is etiologically heterogeneous and that basaloid or warty SCC (BWSCC) and vulvar intraepithelial neoplasia (VIN) are linked to human papillomavirus (HPV) infections while keratinizing SCC (KSCC) is a non-HPV-associated malignancy. In the present study, HPV-specific antibodies in sera of patients with BWSCC, VIN, and KSCC and of controls were examined by ELISA for antibodies reactive to HPV-16 virus-like particles (VLP) and in radioimmunoprecipitation assays for antibodies to HPV-16 E6 and E7 proteins expressed by in vitro transcription and translation. The prevalences of antibodies to HPV-16 VLPs were significantly higher in HPV-associated VIN (59.1%) and BWSCC (50.0%) than in KSCC (22.2%) and controls (18.2%). Antibodies to E6 and E7 proteins were more prevalent in BWSCC than in any other groups. Prevalence of serum antibodies to any one of the antigen preparations was significantly higher in BWSCC (64.3%) and VIN (59.1%) than in KSCC (27.8%) and controls (22.2%). Also, sera with high antibody titers were found more frequently in BWSCC and VIN cases than in controls. These data provide immunological evidence in support of the observation that VIN and BWSCC, but not KSCC, are associated with HPV infections.