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Factors Affecting the Stability and Accuracy of the Bioassay for the Sperm Attractant Sirenin ¹

^a Department of Botany, University of California, Berkeley, California 94720

Optimal response of the sperm of Allomyces from the highly male strain M16 to the chemotactic agent, sirenin, was shown to occur when the sperm suspension contained 2 mM piparazine-N', N-bis[2-ethane sulfonic acid] buffer, 3 mM CaCl₂, and chelated trace elements. For the male strain M3, the CaCl₂ needed was 3.5 mM with the other two components the same as for M16. The inclusion in the sperm suspension of MgCl₂, KH₂PO₄, or NH₄Cl was without effect, except that under certain conditions phosphate was detrimental. The variability of 10 replicate assays was substantially reduced by using sperm in the bioassay at a concentration of 500,000 per ml rather than the former concentration of 100,000 per ml with a concomitant reduction in the concentration of sirenin above the membrane to which the sperm were attracted.