It is well established that water quality is directly linked to health. In-line chlorination is one technology that can be used in the developing world to potentially inactivate pathogens and improve water quality. The purpose of this study was to determine the effectiveness of the Panamanian Ministry of Health's in-line PVC chlorinator under three different operating conditions in a rural water supply system. Free and total chlorine were measured entering the storage tank, leaving the storage tank, and at three households along the transmission line of the water system in the two rural indigenous communities of Calabazal and Quebrada Mina in western Panama during April-August 2011. The Ct method for disinfection was used to compare the measured free chlorine concentration to the concentration required to inactivate common pathogens found in gravity flow water systems in Panama, such as E. coli, Salmonella typhi, Hepatitis A, Giardia lamblia, and E. histolytica, as well as other pathogens of interest to the global health community, such as Vibrio cholerae and Rotavirus. When the chlorine tablet was sealed in a plastic wrapper prior to use to prevent contact with humid surroundings, the chlorine was able to dissolve in seven days instead of three hours into the transmission line. The use of one tablet, sealed in a plastic wrapper before use, was able to obtain the required free chlorine concentration estimated to disinfect E. coli, Vibrio cholerae, Rotavirus, Salmonella typhi, and Hepatitis A. However, it did not achieve a free chlorine concentration above 0.27 mg/L needed to inactivate Giardia lamblia nor above 0.35 mg/L needed to inactivate E. histolytica. The use of three properly stored tablets in the chlorinator was able to provide a free chlorine concentration above 0.35 mg/L for only one day, reaching 0.37 mg/L, before falling below 0.35 mg/L to a level of 0.26 mg/L the next day. The study suggests that with three tablets the in-line PVC chlorinator can be an effective technology if slightly more free chlorine concentration can enter the system. The cost of this technology could be allocated to every owner with a house connection in the communities of Calabazal and Quebrada Mina by increasing their monthly tariff by $1 each month.