A prospective study was conducted in 4339 HIV-positive clients at the Thai Red Cross AIDS Research Centre (TRC-ARC) Anonymous clinic, Bangkok, Thailand between January 2003 and April 2008. A tuberculin skin test (TST) was done for all patients without a previous history of tuberculosis (TB). Nine months of isoniazid (INH) was given for all positive TST/no active TB. TST-negative clients were asked to repeat the TST annually. The study aim was to evaluate the feasibility and efficacy of INH prophylaxis for preventing TB in HIV-positive Thai patients. Of those patients, 4111 (94.7%) had a TST done; 1157 (28.1%) were TST positive and 799 patients started INH prophylaxis. In all, 551 (69%) and 633 (79.2%) patients completed 9 months and 6 months of INH, respectively; 176 (20.2%) patients had a negative TST at baseline and subsequently converted to positive. Only patients with a baseline CD4 >200 cells/μL (p=0.000) and currently on antiretroviral (ARV) treatment (p=0.000) were related to having a positive TST. This baseline CD4 level was also significantly related to higher INH completion rates at 6 months (p=0.000). Interestingly, none of INH completion patients developed active TB. The feasibility of INH prophylaxis in TST-positive patients in this setting is possible. However, the long-term advantage of INH prophylaxis in terms of TB prevention, especially in HIV-1-infected patients on highly active antiretroviral therapy (HAART), is still an issue that needs more research.