524 FEANK R. LILLIE 



a second mixing up, and only to a slight extent or not at all after 

 a third mixing. This condition of relative inactivity, or staleness, 

 is reached in a few minutes, but varies more or less according to the 

 density of the suspension, a very dense suspension exhibiting it 

 more quickly than one less dense. The activity of the sperm 

 may be restored, partly at least, by the addition of fresh sea- 

 water, which shows that the staleness is not due to exhaustion, 

 l)ut to the accumulation of by-products of activity in the sea- 

 ^^•at{u•. Of these the chief is probably CO2, as will be shown by 

 (^xj)eriments described beyond. Tlie formation of CO2 by the 

 acti\'ity of the spermatozoa themselves is indeed one of the chief 

 causes that limits their activity when sufficiently concentrated to 

 form milky suspen.sions. To obtain the best results with the 

 experiments described it is necessary to work with fresh sperm; 

 otherwise, the accumulation of CO2 may obscure the reactions. 



4. Temperature 



In 1911 a series of observations were made on the effect of tem- 

 perature on the aggregation reaction of fresh sperm. In general 

 the results as tabulated are: 



13°C. No aggregations form 



15°C. Slight signs of aggregation in 4 minutes 



18°-19°C. Aggregation in from 2 to 4 minutes; much fewer in number than at 

 higher temperatures 



20.r)°C. Aggregations, numerous, in 1 minute 



23.5°C. Aggregations, yet more numerous, in 30 seconds 



2().r)°C. No aggregations form at this temperature, but they form as the tem- 

 perature falls to 23°C. 



In general temperatvu-es from 20° to 23.5°C. are optimum for 

 the aggregation phenomenon. At 15°C. the movements of the 

 spermatozoa are too slow, and at 26.5°C. the movements are 

 extremely active, but apparently uncoordinated, so that the aggre- 

 gation reaction is not given. These figures possess no absolute 

 value, but they indicate approximateh^ the limits of temperature 

 within which th(^ reaction may be expected. The normal tem- 

 perature of t he s(>a-water \'ari(^s from about 18° to 22°C. at Woods 

 Hole during the In'eeding season. 



