The ideal microbicide or microbicide combination must have activity against cell-free and cell-associated primary HIV isolates representing multiple clades, must inhibit transmission to relevant cell types within the mucosa, and must retain activity in the presence of cervicovaginal fluid and when virus is introduced in semen or seminal plasma. This summary briefly reviews some of the basic concepts underlying the optimization of assays to evaluate the efficacy of candidate microbicides against HIV/STI. The review is based on the presentations of Drs. Stuart Turville (Population Council), Dan Barouch (Harvard University), Scott McCoombe (Northwestern University), Betsy Herold (Albert Einstein College of Medicine), Sam Niedbala (Lehigh University), and the subsequent discussion led by Drs. Robin Shattock (St. George's Hospital Medical School) and Jim Turpin (National Institutes of Health) during a session entitled "Biomarkers of HIV/STI" held at the conference entitled "Biomarkers for evaluation of vaginal microbicides and contraceptives: discovery and early validation," organized by CONRAD and the Alliance for Microbicide Development in November of 2006 as well as more recent published findings.