FVIII is more immunogenic than is OVA. FVIII^−/−/BALB/c mice received 5 weekly injections of 1 μg FVIII intraperitoneally (n = 6) or 1 μg OVA intraperitoneally (n = 6). This experiment has been repeated at least 5 times, and representative results are reported here. One week after the final injection, peripheral blood was collected for ELISA analysis and splenocytes were stimulated in vitro for proliferation analysis. (A) Serum samples were assayed on ELISA plates that had been coated with either FVIII or OVA. To compare responses to different proteins, the endpoint dilution is calculated as the highest dilution that maintains a reading twice background. The immunogens are indicated on the x-axis and the titers versus the same antigen on the y-axis. The mean values for the mice treated with OVA (308 ± 106) were more than 2 logs decreased from the mice treated with FVIII (53 000 ± 23 000). (B) Splenocytes were processed into a single-cell suspension and assayed in a thymidine (³H) incorporation assay described previously. T cells from FVIII-treated mice proliferated in response to coculture with FVIII antigen, whereas cells from OVA-treated mice respond poorly to OVA. The background counts (∼ 2000) have been subtracted (Δ counts), and the difference between the groups was highly significant (**P = .004).