Cell-free HTLV-I particles mediate in vivo infectivity. (A) HTLV-I provirus was detected in rabbit PBMC; (B) HTLV-I antibodies in rabbit sera were detected using a western blot assay (Genelabs Techologies, Singapore). A goat anti-rabbit IgG conjugated with alkaline phosphatase (Santa Cruz Biotechnology, Santa Cruz, CA) was used for rabbit samples instead of goat anti-human IgG conjugate provided by the kit. Mo., month post inoculation; +, positive control serum; -, negative control serum; rgp46, HTLV-I envelope recombinant protein; gp46, HTLV-I env protein; p19 and p24, HTLV-I gag proteins; GD21 specific HTLV-I and HTLV-II epitope recombinant envelop protein. (C) HTLV-I gag p19 protein was detected in the culture supernatants of rabbit PBMC taken at one month post inoculation. (D) Schematic representation showing the regions sequenced. The target regions were amplified by PCR and purified PCR products served as templates for direct sequencing. Stable transmissions of HTLV-I sequence fragments in rabbit BH24, TO11, TO12 and BH42 were observed. No mutation was detected in the analyzed LTR, gag, pol, env, and rex regions for the period of observation up to 20 months. The red arrows indicate the primers used to amplify an env fragment in real-time QPCR assay.