This study was designed to determine the effects of stage of lactation and subsequent energy status on metabolic and endocrine measures, follicular development, and the quality of oocytes obtained from Holstein cows. Holstein cows were selected prior to calving and assigned to the early lactation (EL) group (n=8) while, cows at d 90 postpartum were selected for the mid-lactation (ML) group (n=7). Blood samples were taken twice weekly from 4 wk prior to the start of follicular aspirations and then on through the aspiration periods for metabolite and hormone determination. Ultrasound-guided transvaginal follicular aspiration (TVFA) was conducted twice weekly for a 10-wk period on all cows. Follicular fluid samples were obtained from the largest follicle, > 10 mm in diameter, for hormone determination. All data were analyzed by ANOVA, using the general linear model procedures. Mean energy balance was positive for (2.43 ± 0.32 Mcal/kg) for ML cows and negative (-1.55 ± 0.33 Mcal/kg) for EL cows. In ML cows serum progesterone (P4) decreased rapidly from 2.7 ± 0.1 ng/ml at the first aspiration session to a nadir of 0.33 ± 0.1 ng/ml at wk 8, while follicular fluid P4 increased from 0.9 ± 0.5 to 5.6 ± 0.5 ng/ml. In the EL cows serum and follicular fluid P4 remained relatively constant over the course of aspirations. There was a linear increase in follicular fluid insulin-like growth factor I (IGF-I) for EL and ML cows, however the increase was more rapid for ML cows (159 ± 36 to 200 ± 36 ng/ml) than for EL cows (145 ± 36 to 164 ± 36 ng/ml). Over the aspiration period nonesterified fatty acids (NEFA) declined rapidly for the EL cows (0.32 ± 0.2 to 0.22 ± 0.2 mEq/L), while serum NEFA for the ML cows were relatively stable (0.19 ± 0.2 to 0.22 ± 0.2 mEq/L). The number of follicles observed during the aspiration sessions increased linearly for both EL and ML cows (P < 0.05) over the 10-wk period. However, the increase was larger for the ML cows than for the EL cows, going from 14.2 ± 0.5 to 18.1 ± 0.5 and 14.9 ± 0.3 to 15.7 ± 0.5, respectively. These results show that cows in early lactation are physiologically under more production stress than cows in mid lactation. Furthermore, increasing levels of serum and follicular fluid IGF-I in mid lactation may reflect differences in follicle and oocyte measures.