p35 and p37 are Borrelia burgdorferi genes encoding 35 and 37 kDa proteins. The gene products were identified by differential screening of a B. burgdorferi expression library with sera from B. burgdorferi infected- and B. burgdorferi-hyperimmunized mice. Northern blot and RT-PCR analyses confirmed that these genes were selectively expressed in vivo. ELISA, using P35 and P37, showed that infected mice (5 of 5, 100%) and patients (31 of 43, 72%) with Lyme borreliosis developed P35 or P37 antibodies. Mice developed peak IgG titers to P35 and P37 within 30 days, followed by decline. Mice given both P35 and P37 antisera were protected from challenge with 10(2) B. burgdorferi, and P35 and P37 antisera also afforded protection when administered 24 hr after spirochete challenge. The use of in vivo-expressed antigens such as P35 and P37 represents a new approach for Lyme disease serodiagnosis and for understanding the role of B. burgdorferi-specific immune responses in host immunity.