1. Comparison of different methods for determining bovine serum Anti-HCG.
The original method to determine bovine serum Anti-HCG reported in our earlier papers was as follows : Two parts (2/3 ml.) of undiluted serum sample and 1 part (1/3 ml.) of saline solution of HCG with graded concentrations were injected into inguinal region of immature female mice subcutaneously twice daily for 2 days, and these animals were killed 100 hours after the first injection, then ovarian responses were observed. In the present study, the original method was compared with two other methods. The first method (Serum dilution method) : Serum sample (2/3 ml.) which was successively diluted by saline and saline solution of HCG (1/3 ml.) with a certain concentration were mixed, and the mixture was assayed in the same way as in the original method. The second method (Simultaneous injection method) : Undiluted serum sample (2/3 ml.) and saline solution of HCG(1/3ml.) which was successively diluted were injected simul-taneously into inguinal and axillary regions of immature female mice respectively. The results of the two experiments were shown in Table 1 and 2, respectively. From these results, no appreciable differences in HCG inhibiting titers between the original method and the two other methods could be found.
2. Does bovine serum Anti-HCG against a certain HCG preparation inhibit gonadotrophic activities of HCG prepared by another maker.
Several workers have shown recently that the antigenicities of gonadotrophin preparations are variable according to their manufacturing processes or grades of purity. In the present experiments, it was examined whether the bovine serum Anti-HCG formed against a certain HCG preparation might inhibit gonadotrophic activities of HCG prepared by another maker or not. Inhibition titers of Anti-HCG positive serum samples obtained from experimental cattle following injections of HCG-A were examined against HCG-A, -B, -C, -D and-E respectively ; HCG-A, -B, -C, -D and-E had been prepared by different maker-A, -B, -C, -D and-E respectively. The determinations were performed according to the serum dilution method mentioned above. The results obtained were shown in Table 3. The results revealed that not only HCG-A but HCG-B, -C, -D and-E were inhibited by bovine serum Anti-HCG-A, the inhibiting titers being nearly equal between them.
3. Examination of inhibitory effects of bovine serum Anti-HCG on gonadotrophins from another origins. In hibitory effects of bovine serum Anti-HCG on gonadotrophic activities of pregnant mare serum gonadotrophin (PMS) and sheep anterior pituitary gonadotrophin (SAPG)were examined according to the original method of Anti-HCG determination. The results obtained were shown in Table 4 and 5 respectively. From these experiments, it was presented that gonadotrophic activities of PMS and SAPG could not be inhibited by bovine serum Anti-HCG. Therefore, it was supposed that application of PMS or SAPG might be effective on treating ovarian disorders of cattle in which Anti-HCG were formed following repeated HCG injections.

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