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A vertical polyacrylamide gradient gel (3% to 7%) was designed to facilitate the electrophoretic resolution and classification of isoenzyme patterns of Entamoeba histolytica isolates. The following enzyme systems were used: phosphoglucomutase (PGM), hexokinase (HX), glucosephosphate isomerase (GPI), and malate dehydrogenase (ME). The modifications in the electrophoretic procedure and sample preparation allowed the reproducible comparison of enzyme patterns of axenic, monoxenic, and mixed cultures of E. histolytica isolated from humans. The clear distinction obtained in gradient polyacrylamide gels, between amebic isoenzyme bands and those from bacteria, renders this technique adequate for application to epidemiological studies where mixed cultures are used. The isoenzyme patterns of eight isolates from asymptomatic carriers, rigorously characterized by the absence of clinical, endoscopic, and serological findings were studied and compared with three well characterized pathogenic strains, cultured under axenic conditions. Our observations confirm the existence of distinct isoenzyme patterns for PGM, HX, and GPI in pathogenic and nonpathogenic strains, and reveal the consistent presence of more than one band for GPI. In addition, a previously undescribed band for GPI with an Rf of 0.64 in a carrier strain was found. The results suggest that while carriers usually harbor amebas with nonpathogenic isoenzyme patterns, pathogenic patterns also may be found in carriers.
Various slight modifications of electrophoretic techniques were tried to obtain more reproducible results for comparing several isozymes with individual mosquito larvae. Horizontal electrophoresis was done using polyacrylamide gel plates of 1 mm thickness on a simple cooling system in which ice-cold water was circulated. This made it possible to carry out electrophoresis effectively even during field work in tropical areas. Gel concentration in routine work is 5% for usual enzymes, but for esterase, 6% gels have been adopted. Gel concentrations, running distance, running time, visualization of isozyme bands, etc., have especially been considered.
Akaline phosphatase activity following polyacrylamide gel electrophoresis of human sera obtained from normal populations in the Eastern India shows a correlation between the presence of p++ phenotypes with the B and O blood groups only in females. The incidence of additional bands in the beta-lipoprotein zone is much higher in female samples than in males. The frequencies of both p++ and p+ henotypes within the different endogamous communities and castes are rather variable. Electorphoresis following heat treatment of sera obtained from pregnant females shows the complete absence of p++ and p+, as also of bands at the beta-lipoprotein zones in them.
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Since detection of the first stage of nephropathies requires an exact definition of physiological proteinuria, this has been sought in 97 healthy individuals. Measured by a modification of the biuret reaction, physiological proteinuria did not exceed 100 mug/min in the recumbent position (average of 24.5 mug/min) and 150 mg/24 h (average of 51 mg/24 h). No significant differences were seen with respect to age or sex. Qualitative analysis of urinary proteins was done by cellulose acetate electrophoresis, immunoelectrophoresis and polyacrylamide-SDS electrophoresis. This latter method has the great advantage of classifying the proteins according to their molecular weight, wtihout electrostatic interference. Its graphic representation delineates a normal zone allowing an objective distinction between physiological and pathological proteinurias. In the majority of cases, the proportion of albumin is between 25 and 55%. The orthostatic position increases proteinuria (average of 41 mug/min) with a tendency to a distribution which erroneously suggests a glomerulopathy. Accordingly, investigation for small changes in proteinuria should always be carried out on urine formed in the recumbent position.
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After previous examinations in normal parotid glands in this paper secretion of proteins in parotid glands with pleomorphic adenomas, Whartin tumors, parotid carcinomas, chronic parotitis and sialadenosis should be studied. Different discelectrophoretic and immunelectrophoretic runs were done. Thereby we got the following changes in electrophoreses: Pleomorphic adenomas generally show a more palid pattern in band-structure, caused by the diminished proteinconcentration in the secretions. Most remarkable are thereby the c1 and d1-4-bands. Immunelectrophoretic separations are normal. Whartin tumors show a different bandpattern in the anodal near gel region, which is enpowered by immunelectrophoretic runs. Secretions from parotid carcinomas show a characteristic change in bandstructure. There are found added bands which result from serumproteins as immunelectrophoretic examinations proof. The same is found in glands with parotitis in contrary to glands with sialadenosis.
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