Cell type- and virus strain- independent restriction of HIV-1 replication by semen exosomes. (A and B) Monocytic cell lines (A) U937 and (B) THP1; (C to F) T lymphocyte cell lines; (C) Rev-CEM, (D) JURKAT, (E) SupT1, (F) PM1, and (G) PBL (from 3 different healthy donors) pretreated ± AZT for 2 h were exposed to HIV-1SF-162 or HIV-1NL4-3 pre-incubated with PBS or SE for 1 h. Total DNA isolated from cells was examined for levels of HIV-1 Gag DNA 24 h later by qPCR. (H) U937, THP1, Rev-CEM, JURKAT, SupT1, and PM1 were exposed to HIV-1SF-162 or HIV-1NL4-3 pre-incubated with PBS or SE for 1 h. Total RNA was examined for levels of HIV-1 Gag RNA 24 h later by RT-qPCR. (I) PBL from 3 healthy donors were pretreated ± AZT for 2 h and exposed to HIV-1NL4-3 pre-incubated with PBS or SE. Infectivity was assessed by qPCR and RT-qPCR for detection of HIV-1 Gag DNA or RNA respectively. (J) Total DNA was examined for integrated HIV-1 DNA in U937 cells exposed to HIV-1SF-162 or SupT1 cells exposed to HIV-1NL4-3 in the presence or absence of SE or AZT by Alu-PCR. (K) TZM-bl cells exposed to R5-monotropic clade B (SF162) or X4-monotropic clade B (IIIB and U1) pre-incubated with PBS or SE. HIV-1 pre-incubated with PBS was set at 1. (L) TZM-bl cells exposed to transmitted/founder (T/F) molecular infectious clones (REJO and CH162) of HIV-1 pre-incubated with PBS or SE were examined for infectivity by luminescence output. HIV-1 pre-incubated with PBS was set at 1. **P < 0.02 and Student's t test was used for all samples. Data are expressed as mean ± SD and presented as fold change of vehicle. Experiments were repeated at least three times with similar results.