On what day of a menstrual cycle should an estrogen and progesterone test be done?

Different days: Estradiol (e2) can be checked as a 'baseline' level on day 2 or 3, or later in the first half of your cycle if you are taking fertility medications - often combined with ultrasound monitoring of your follicles. Progesterone is typically checked on day 21 if you have 28 day cycles, or a week after ovulation if your cycles are longer or shorter. Progesterone can also be checked when you're pregnant.

21st Day: In order to know whether ovulation occurs, there should also be a mid-luteal Progesterone blood test on cycle days 20-22.