The efficiency of an alternative instrument-free testing strategy was evaluated using a membrane-based rapid screening assay (HIVCHEK and its new version HIVCHEK1 + 2) in serial combination with a particle agglutination assay (SERODIA-HIV). Among 1,054 Zairian individuals at high risk of HIV infection, 573 were Western blot-positive for HIV-1 (54.4%) and none were Western blot-positive for HIV-2. In this group, the sensitivities of the serial combination HIVCHEK plus SERODIA-HIV and HIVCHEK1 + 2 plus SERODIA-HIV were 98.1 and 98.2%, respectively, and the specificities were 99.6 and 99.5% compared with HIV-1 Western blot. The positive predictive values were 99.6% for HIVCHEK plus SERODIA-HIV and 99.5% for HIVCHEK1 + 2 plus SERODIA-HIV; the negative predictive values were 97.8 and 97.9%, respectively. Among 1,495 pregnant women, 90 were Western blot-positive for HIV-1 (6.0%), and 54 of 1,510 blood donors were HIV-1 Western blot-positive (3.6%). None were positive for HIV-2. The sensitivities of HIVCHEK plus SERODIA-HIV and HIVCHEK1 + 2 plus SERODIA-HIV in these groups were 98.6 and 99.3%, respectively, and the specificities were 99.8 and 99.7%. The positive and negative predictive values of HIVCHEK plus SERODIA-HIV were 96.6 and 99.9%, respectively, and they were 94.1 and 99.9%, respectively, for HIVCHEK1 + 2 plus SERODIA-HIV. These instrument-free testing strategies are efficient alternatives for serodiagnosis of HIV-1 infection, although their cost should be further reduced.