Homogenates of the mosquitoes, Aedes aegypti, Anopheles quadrimaculatus, and Culex quinquefasciatus contain aldolase systems which are quite similar in physical characteristics. The apparent Michaelis-Menten constants ranged from 3.2 X 10-3 M to 3.6 X 10-3 M, and the optimal pH's lay between pH 7.4 and pH 8.2. The optimum temperature for Anopheles aldolase was about 20 C, whereas that for both Culex and Aedes aldolases was approximately 30 C. The sex of the mosquito had no effect upon the aldolase activity. The aldolase activity of each species was enhanced by ethylenediaminetetraacetate and was inhibited by the cations magnesium, manganese, and calcium. Although studies have been carried out on the general oxidative metabolism of mosquitoes (Gonda et al., 1957; Mercado, Trembley, and von Brand, 1956; Tekle, 1960), relatively little information is available on the intermediary glycolytic metabolism of mosquitoes and virtually none concerning characterization of the specific enzymes of this cycle. Several studies on the characteristics of aldolase in organisms other than mosquitoes have been published. Bard and Gunsalus (1950) in a study of this enzyme in Clostridium perfringens determined the physical conditions necessary for optimal activity. Baerstein and Rees (1952) described the aldolase from culture forms of Trypanosoma cruzi, and Baernstein (1955) followed this with a study of this enzyme in the culture form of Trichomonas vaginalis. Warburg and Christian (1943) studied the properties of yeast aldolase, and, in 1940, Herbert et al., published a definitive survey of the physical properties of rabbit muscle aldolase. A helminth aldolase, that of Taenia crassiceps, was described by Phifer in 1958. This paper is concerned with the characterization and comparison of aldolase in homogenates of three mosquito genera-Aedes aegypti, Anopheles quadrimaculatus, and Culex quinquefasciatus, including the determination of conditions for optimal activity and the effects of sex and of metal ions. Received for publication 29 January 1962. * Department of Health, Education, and Welfare, Public Health Service, National Institute of Allergy and Infectious Diseases, Laboratory of Parasite Chemotherapy, Section on Epidemiology, P. O. Box 717, Columbia, South Carolina. MATERIALS AND METHODS The mosquitoes used were strains of Aedes aegypti, Anopheles quadrimaculatus, and Culex quinquefasciatus which have been maintained in the laboratory for several years. When the mosquitoes were 5 to 6 days of age, they were frozen at -70 C for storage. Two hundred milligrams of each species were homogenized in 15 ml of tris (hydroxymethyl )aminomethane (0.1 M) buffered at pH 8.6. Fructose diphosphate (FDP), used as substrate, was a commercial preparation of the sodium salt. The reaction mixture was essentially that described by Sibley and Lehninger (1949). The reaction mixture contained: tris( hydroxymethyl) aminomethane (tris) 0.1 M, pH 8.6; hydrazine 0.056 M, pH 8.6; fructose diphosphate, pH 8.6 at various concentrations, ranging from 0.0009 M to 0.0479 M; homogenate in tris buffered at pH 8.6; and water to make 5 ml. Various additional substances such as MgSO4 and FeCl3 were substituted for an equal volume of water. Tubes were equilibrated 6 minutes in the 37 C water bath before homogenate was added. A blank was prepared by adding 4 ml of 10 per cent trichloracetic acid (TCA) to one tube for each experimental variable before the homogenate was added. After 15 minutes incubation, TCA was added to each of the other tubes and the mixtures were centrifuged. The samples were then handled according to the Sibley and Lehninger procedure and color density was measured in the Bausch and Lomb colorimeter at a wave length of 540 millimicrons. The quantity of dinitrophenylhydrazone was calibrated in order that the results might be expressed in terms of alkali-labile phosphate. The technique of Fiske and SubbaRow (1925) as modified by Horwitt (1952) was used. In the data presented below, enzymatic activity is expressed in micrograms of alkali-labile-phosphate (ALP) per hour. It is recognized that mosquito homogenates contain a small number of bacteria, but the numbers involved would not significantly alter enzyme activity.