Rapid detection of 6-phosphogluconate dehydrogenase variants by electrophoresis on cellulose acetate gel

A procedure, suitable even under field conditions for the rapid detection of 6-phosphogluconate dehydrogenase electrophoretic variants, is described.Work supported by N.I.H.—grant no. ROI GM 13415.     

Differentiation of mite species by using cellulose acetate and equilibrium polyacrylamide gel electrophoresis of isoenzymes

The isoenzymes of various species of medically and economically important mites were studied using cellulose acetate and equilibrium polyacrylamide gel electrophoresis. Interspecific differences in isoenzymes were found, which were species-specific. Intraspecific differences in isoenzymes were also found when individual mites were examined. The efficiency of each technique, and their use in various fields of acarology, are discussed. In addition, possible phylogenetic relationships as revealed by these techniques are suggested.     

Diagnostic performance of lactate dehydrogenase (LDH) isoenzymes levels for the severity of COVID-19

BackgroundLactate dehydrogenase (LDH) levels predict coronavirus disease 2019 (COVID-19) severity. We investigated LDH isoenzyme levels to identify the tissue responsible for serum LDH elevation in patients with COVID-19.MethodsHospitalised COVID-19 patients with serum LDH levels exceeding the upper reference limit included. LDH isoenzymes were detected quantitatively on agarose gels. The radiological severity of lung involvement on computed tomography was scored as 0-5 for each lobe (total possible score, 0-25). Disease severity was determined using the World Health Organization (WHO) clinical progression scale.ResultsIn total, 111 patients (mean age, 59.96 ± 16.14), including 43 females (38.7%), were enrolled. The serum levels of total LDH and all five LDH isoenzymes were significantly higher in the severe group. The levels of all LDH isoenzymes excluding LDH5 positively correlated with the WHO score. LDH3 levels correlated with chest computed tomography findings (r² = 0.267, p = 0.005). On multivariate analysis, LDH3 was an independent risk factor for the deterioration of COVID-19.ConclusionsLDH3 appears to be an independent risk factor for deterioration in patients with COVID-19. LDH elevation in patients with COVID-19 predominantly resulted from lung, liver and muscle damage.     

Additional lactate dehydrogenase (LDH) isoenzymes in normal testis and spermatozoa of adult man

Summary Adult human testicular tissue contains up to six previously undescribed lactate dehydrogenase (LDH) isoenzymes in addition to the five LDH isoenzymes normally found and the sixth found in spermatogenic cells and spermatozoa, LDH-X. Additional LDH isoenzymes were also found in spermatozoa but not in seminal fluid or in serum. After electrophoresis one additional LDH isoenzyme of testicular tissue was localized between LDH-1 and LDH-2, two between LDH-2 and LDH-3, two between LDH-3 and LDH-4, and two between LDH-4 and LDH-5. These localizations indicate that the additional LDH isoenzymes are tetramers combining the A and B subunits of the five normal LDH isoenzymes and the C subunit of LDH-X. The additional LDH isoenzymes may be important in the metabolism of spermatogenic germ cells and spermatozoa.     

Esterase-D polymorphism in Assam by cellulose acetate electrophoresis

Summary With the help of a simplifed and quick method, cellulose acetate electrophoresis, the phenotypes of esterase D were determined in an Assamese population. The gene frequencies of Es D¹ were 0.7263 and 0.2737 for Es D².

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Zusammenfassung In einer Stichprobe aus Assam wurde mit Hilfe einer einfachen und schnellen Methode, der Cellulose-Acetat-Elektrophorese, die Bestimmung der Esterase D-Phänotypen durchgeführt. Die Genfrequenzen wurden für Es D¹ zu 0,.7263 und für Es D² zu 0.2737 bestimmt.

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Supported by the Deutsche Forschungsgemeinschaft, the Stiftung VW and the Fonds der Chemischen Industries.     

Two-dimensional electrophoresis on the layers of cellulose acetate membrane

A new type of two-dimensional electrophoresis for analysis of protein using cellulose acetate membrane has been developed. Prior to the separation, proteins in a sample are concentrated to a narrow zone on a strip of cellulose acetate according to “steady-state stacking” of isotachophoresis. Electroendosmotic counterflow on cellulose acetate membranes is advantageous for the isotachophoretic concentration of large sample volumes. The concentrated protein zone is then subjected to electrophoretic separation on the same strip. This first-dimensional separation including the concentrating process is named “concentrating electrophoresis.” Iso-electric focusing on several layers of cellulose acetate membrane is performed in the second-dimensional step. Many kinds of detection methods can be applied to the layers among which proteins are distributed. The novel two-dimensional electrophoresis takes only 5 h to perform.     

Beta-agonists enhance the lactic dehydrogenase (LDH) expression in serum and ventricular myocytes of mice

Beta-agonists have skeletal muscle specific protein anabolic effects and are also known to cause cardiac hypertrophy. Changed total LDH and its isozymic patterns are conveniently employed for the detection of different pathophysiological states of the tissues. The purpose of this study is to confirm total LDH and its isozymic expression in ventricular tissue and serum in mice following oral administration of single but higher dose of isoproterenol (Iso) and clenbuterol (Cl) (100 mg/kg body wt. and 20 mg/kg body wt., respectively), after 4, 8 and 20 hours of drug administration. Mice heart witnessed increased total LDH levels with time. Serum on the other hand showed decline in total LDH concentrations at the initial points of the drug treatment. No doubt, total LDH expression increased towards 20th h post-drug treatment but this increase is mainly due to anaerobic isozymes, i.e. LDH4 and LDH5. The findings of the present study suggest that tissue damage is definitely caused by two beta-agonists after giving single dose for shorter time span (20 hours) and the impact of the damage varies from drug to drug. Increase in total LDH in serum is not due to release from heart but from some other tissues having anaerobic metabolism.     

Subunit homologies of lactate dehydrogenase (LDH) isoenzymes between amphibians (Xenopus laevis) and mammals (Wistar rat)

• 1.1. The subunit distribution and subunit homologies of LDH isoenzymes were studied in the amphibian Xenopus laevis and in Wistar rats.
• 2.2. Several of the 11–15 isoenzymes of the pattern in Xenopus, separable by vertical starch gel electrophoresis, were purified, hybridized, and the cross-reaction of antibodies against the most positively charged isoenzyme with the isoenzymes present in tissue extracts of both species was tested.
• 3.3. The isoenzyme with the highest positive charge in Xenopus is a M-homotetramer homologous to mammalian LDH₅.
• 4.4. The multibanded pattern of Xenopus LDH isoenzymes is very probably due to heterozygoty of the gene locus controlling the synthesis of M-subunits rather than to an epigenetic subbanding phenomenon.

     

Production of monoclonal antibodies to lactate dehydrogenase (LDH) isoenzymes for immunohistochemical study on fixed tissue section

Summary Purified lactate dehydrogenase (LDH) isoenzyme 1 (H or B subunits) and isoenzyme 5 (M or A subunits) were used to prepare monoclonal antibodies (MAb) suitable for immunohistochemical detection on formalin fixed paraffin-embedded tissue sections. In the initial fusions, screening of the antibodies was based on enzyme linked immunosorbent assay (ELISA) against the immunogens. None of the antibodies obtained was satisfactory. There were various problems related to specificity, crossreactivity, affinity and also the properties of the monoclonal antibody itself. Using a combined system involving more than one method for screening, two suitable monoclonal antibodies, MAb65 (to H-type LDH) and MAb25 (to M-type LDH) were selected. Both antibodies reacted specifically with corresponding LDH isoenzymes as shown in a series of tests. Their reactivity in sections of formalin fixed paraffin-embedded tissue indicated that both antibodies are suitable reagents for immunohistochemical studies.