We assessed the possible effects of the volatile halogenated anesthetics halothane, enflurane, and isoflurane on Ca 2ุ electrode measurements and on the Ca 2ุ sensitivity of the bioluminescent protein aequorin. In Ca 2ุ -EGTA buffers of different pCa values (7.870, 6.726, 6.033, 4.974, 4.038, and 2.995) and in serial Ca 2ุ dilutions (10 ุ4 , 10 ุ3 , and 10 ุ2 M), halothane, enflurane, and isoflurane each caused a concentration-dependent and reversible increase in the absolute value of the negative electrode potential. Isoflurane and enflurane had larger effects than halothane. Neither of these anesthetics changed aequorin luminescence at any pCa tested in the range 2-8. There was no potentiation or inactivation of aequorin luminescence over a period of up to 2 h. These results suggest that (1) halothane, enflurane, and isoflurane interfere with Ca 2ุ electrode measurements, most likely by changing the physicochemical properties of the membrane; (2) these anesthetics do not inactivate or otherwise modify the characteristics of the reaction of Ca 2ุ with aequorin; and (3) these anesthetics do not change the apparent affinity of EGTA for Ca 2ุ .