Plasma androstenedione (A) and testosterone (T) were measured in 47 prepubertal and pubertal boys ages 4–15 yr 9 months using 35S-thiosemicarbazide double isotope methods. Before age 11 plasma A was 62±8.8 (se) ng/100 ml, while T was 43.7±5.7 (se) ng/100 ml. Values are corrected for the mean blank of the respective methods. After age 11 there was a variable rise in the plasma concentration of T. Prepubertal values for T (<100 ng/100 ml) were found in 1/3 of boys ages 11–15 yr 9 months. Plasma A did not change with chronological age. Plasma T values were grouped by pubertal stage as defined by pubic hair and genital development (Tanner). Mean plasma T in stage I–IV was 39±5, 73±19, 182±55 and 465±66 (se) ng/100 ml, respectively. Only a few pubertal values for plasma T were within the normal adult male range, suggesting that puberty is associated with an extended period of increasing testosterone secretion. Plasma A did not increase with advancing pubertal development. The administration of gonadotropin to 2 prepubertal boys resulted in an increase in plasma T concentration to pubertal levels. This suggests that interstitial cell maturation is not a limiting factor in the initiation of puberty.