We developed an improved method for accurately measuring telomere lengths based on two-dimensional calibration of DNA sizes combined with pulsed field electrophoresis and quantitative analysis of high-resolution gel images. This method was used to quantify the length of telomeres in longitudinal samples of peripheral blood mononuclear cells (PBMCs) from five chimpanzees infected with human immunodeficiency virus type 1 (HIV-1) and three uninfected animals, 14 to 27 years of age. The average length of the telomere restriction fragments (TRF) of infected and uninfected chimpanzees were 11.7 ± 0.25 kbp, and 11.6 ± 0.61 kbp, respectively, and were about 1 kbp and 3 kbp longer than those of human infants and 30 year old adults, respectively. There was a trend of a slight decrease (30-60 bp per year) in the TRF of two HIV infected chimpanzees over 30-35 months, while the TRF of one naive chimpanzee slightly increased over 20 months. Although the number of chimpanzees in this study is small and no statistically significant linear dependencies on time were observed, it appears that in chimpanzees, rates of shortening of the TRF are comparable or smaller than in adult humans and are not significantly affected by HIV-1 infection, which may be related to the inability of HIV-1 to cause disease in these animals.