. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. AMKBOCYTK A< i( iRK( iATION IN LIMULl'S 553 the addition of amebocytes to the cuvette. During the plateau period, the recorder pen often oscillates with excursions of varying amplitude, presumably produced In- free aggregates passing through the light path. It was determined that the reproducibilitv of aggregation measurements is crit- ically dependent on the temperature and the speed of stirring. Optimal aggregation occurs at 15° C and this temperature is routinely employed for measurement of amebocyte aggregation. Lower
. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. AMKBOCYTK A< i( iRK( iATION IN LIMULl'S 553 the addition of amebocytes to the cuvette. During the plateau period, the recorder pen often oscillates with excursions of varying amplitude, presumably produced In- free aggregates passing through the light path. It was determined that the reproducibilitv of aggregation measurements is crit- ically dependent on the temperature and the speed of stirring. Optimal aggregation occurs at 15° C and this temperature is routinely employed for measurement of amebocyte aggregation. Lower temperatures (10° C. 5° C, and 0° C) produce progressive inhibition of both the rate and extent of aggregation measured over 2 minutes. Microscopic examination of amebocytes at 0° C shows the presence of aggregates ranging in si/.e from approximately 2-50 cells. Amebocytes within these aggregates retain their individual identity and pseudopods are only occa- sionally seen at the periphery of the aggregates. Stirring speed has a biphasic effect on aggregation. Low speed stirring (60-300 rpm) enhances the rate of. FKH'KK 3. Photomicrographs of amebocytes 2 minutes after withdrawal into a, KDTA ; h, saline ( Nomarski Optics ). aggregation while higher stirring speeds (335—1-50 rpm) appear to inhibit aggrega- tion. A stirring speed of 300-335 rpm is routinely employed. Variation in the number of amebocytes within that range suitable for photometric studies (700- 1600 cells/mnr ) has no effect on the pattern of aggregation. Inhibition oj atnebocyte ut/i/n'</otn»i bv EDTA Buffered EDTA at final concentrations of 17 ± 6 HIM completely inhibits in- creases in transmission over the first two minutes (Fig. 2). In 8 -~ 3 mM EDTA. inhibition of aggregation is significantly decreased and when the concentration of EDTA is reduced to 2 ± 1 mM, inhibition is negligible. Buffered sodium citrate at the same concentrations is less effective than EDTA, but inhibition of aggrega- tion
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Keywords: ., bookauthorlilliefrankrat, booksubjectbiology, booksubjectzoology
