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Six mature cyclic ewes were immunized against an oestradiol-17 β-6-oxo-carboxy-oxime-bovine serum albumin conjugate (oestradiol–protein conjugate). Injection of the conjugate began on day 2 of an oestrous cycle, and subsequent booster injections were given on days 12, 22, 32, 62 and 102.

Six control ewes treated with carrier emulsion alone continued to display regular oestrous cycles, whereas the six immunized ewes failed to return to oestrus after the initiation of treatment. The six immunized ewes failed to ovulate by day 22 of the oestrous cycle in which treatment began, and the preovulatory surge of LH did not occur, although the basal concentration of LH in the jugular plasma rose. The luteal phase of the oestrous cycle in which the first injection of the conjugate was made was normal and three out of six immunized ewes showed typical changes in the cervical mucus at the time of the next expected oestrus. The capacity of the plasma to bind oestradiol-17β rose above the control level after the first booster injection and continued to increase until the end of the experiment, 118 days after the initial treatment. The only significant cross-reactions of the antisera were with oestrone.

The concentration of LH in the plasma rose in parallel with the antisera titres and towards the end of the experiment, a definite episodic pattern of LH secretion was indicated. In immunized ewes the concentration of oestradiol-17β in the plasma increased, reaching 286 ± 36 (S.E.M.) pg/ml, a level far in excess of that found in the normal cyclic ewe. The concentration of progesterone in the circulation was raised for a variable period of time in all immunized ewes between days 30 and 118 after the initial treatment.