Determination Of Lactate Dehydrogenase Isoenzymes Research Articles

Microchip‐based capillary electrophoresis for determination of lactate dehydrogenase isoenzymes

This article describes a novel microchip-based capillary electrophoresis and oncolumn enzymatic reaction analysis protocol for lactate dehydrogenase (LDH) isoenzymes with a home-made xenon lamp-induced fluorescence detection system. A microchip integrated with a temperature-control unit is designed and fabricated for low-temperature electrophoretic separation of LDH isoenzymes, optimal enzyme reaction temperature control, and product detection. A four-step operation and temperature control are employed for the determination of LDH activity by on-chip monitoring of the amount of incubation product of NADH during the fixed incubation period and at a fixed temperature. Experiments on the determination of LDH standard sample and serum LDH isoenzymes from a healthy adult donor are carried out. The results are comparable with those obtained by conventional CE. Shorter analysis times and a more stable and lower background baseline can be achieved. The efficient separation of different LDH forms indicates the potential of microfluidic devices for isoenzyme assay.

Determination of lactate dehydrogenase isoenzymes in single rat glioma cells by capillary electrophoresis with electrochemical detection

A method for determination of lactate dehydrogenase (LDH) isoenzymes in single rat glioma cells (C6) was developed. In this method, a whole cell was electrokinetically injected into the front end of the separation capillary. After that, the cell was lysed by ultrasonication and the isoenzymes in the cell were pre-separated at 20 kV for 5 min and then incubated for 2 min with the enzyme substrates nicotinamide adenine dinucleotide (NAD(+)) and lactate in the capillary electrophoresis running buffer. The electroactive product NADH generated by the isoenzymes through on-capillary enzyme-catalyzed reaction was detected at the outlet of capillary by using the end-capillary amperometric detection with a constant potential mode at a carbon fiber bundle microdisk electrode. Since the amplification of signal via the enzyme reaction, the concentration of nicotinamide adenine dinucleotide (NADH) is much higher than that of LDH. The external standardization was used to quantify isoenzymes in individual cells. Three LDH isoenzymes in single rat glioma cells (C6) were determined and quantified.

Determination of lactate dehydrogenase isoenzymes in single lymphocytes from normal and leukemia cell lines

This work demonstrates that our previously developed technique for single-erythrocyte analysis by capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) can be applied to study individual lymphocytes, with some modification in the cell lysing procedure. A tesla coil was shown to be capable of lysing the lymphocyte cells inside the capillary. The electromagnetic field induced by the tesla coil was believed to be responsible for breaking the cell membrane. The lactate dehydrogenase (LDH) isoenzyme activities and the relative ratios between different LDH isoenzymes were measured for normal lymphocytes as well as B-type and T-type acute lymphoblastic leukemia cells. Both the LDH activity and the isoenzyme ratios show large variations among individual cells. The former is expected due to variations in cell size. The latter implies that single-cell measurements are less useful than the average values over a cell population as markers for leukemia.

Microquantitative determination of lactate dehydrogenase isoenzymes in the myocardium and the conducting system

A newly developed technique was used for the electrophoretic separation of lactate dehydrogenase (LDH) isoenzymes from lyophilized tissue samples in the nanogram range. In this study portions of 10-200 ng from the myocardium and the conducting system of cattle, sheep, pig and man were microdissected and analysed. In the heart tissues of cattle, sheep and pig, the isoforms LDH1, LDH2 and LDH3 were detected in species-specific varying amounts. In all these animals, the conducting system is marked by high LDH1 activity, which is present at a ratio of about 2:1 compared with the myocardium. The values in man, however, differ from these values, but this might be due to post-mortem changes. The findings are discussed with respect to possible aerobic-anaerobic functions.

Ultrathin-layer microelectrophoretic determination of lactate dehydrogenase isoenzymes in corneal and conjunctival epithelium of the cow.

In order to evaluate the impact of tissue oxygenation on the distribution pattern of lactate dehydrogenase isoenzymes, activities of the isoenzymes were measured in microdissected samples of bovine tissue. A highly sensitive ultrathin-layer electrophoretic technique was used to determine the distribution pattern of lactate dehydrogenase isoenzymes in basal, intermediate and superficial layers of the epithelium of central and peripheral cornea and in the epithelium of the bulbar conjunctiva. Measurements revealed almost homogeneous intraepithelial distribution patterns of lactate dehydrogenase isoenzymes in both tissues. In the cornea the lactate dehydrogenase isoenzymes 4 and 5, which are regarded to be specialized for anaerobic glucose metabolism, were found to predominate. In the well-oxygenated conjunctival epithelium most of the activity could be ascribed to the lactate dehydrogenase isoenzyme 3. In contrast to the isoenzymatic activities, total activity of lactate dehydrogenase was inhomogeneously distributed; maximum activities were found in the basal layer of corneal epithelium and in the intermediate layer of conjunctival epithelium. The results indicate that oxygen supply is relevant rather for the intraepithelial distribution of total enzyme activity than for the expression of lactate dehydrogenase isoenzymes.

The diagnostic significance of lactate dehydrogenase isoenzymes in urinary cytology

Lactate dehydrogenase (LDH) isoenzyme distribution was examined in 106 urine samples being tested cytologically for evidence of bladder cancer; the samples were selected to have less than 20 leucocytes and erythrocytes per high power field and the LDH pattern determined by electrophoresis. The Papanicolaou stained-smears showed 68 negative, 17 suspicious and 21 positive. The LDH M-fraction of the urinary supernatant in cytologically positive cases was significantly greater than in negative cases, although the latter included a few false negative samples. Some of the false negatives gave positive results for the LDH M-fraction; these results suggest that the determination of LDH isoenzymes in the urine is useful in diagnosing urinary tract cancers, including early stage, and for follow-up of patients with bladder cancers after surgical resection.

Evaluation of determination of lactate dehydrogenase isoenzyme 1 by chemical inhibition with perchlorate or with 1,6-hexanediol

We have evaluated the determination of lactate dehydrogenase (EC 1.1.1.27) isoenzyme 1 activity by chemical inhibition of the other isoenzymes with perchlorate and with 1,6-hexanediol. In the hexanediol method, we studied the effect of the duration of incubation with the inhibitor; a 5-min incubation yielded results closest to those of an immunochemical technique (Isomune-LD). The perchlorate method was the most precise, and the hexanediol method the least, although for none of the techniques did the coefficient of variation exceed the medically acceptable limit prescribed by the College of American Pathologists. Pairwise correlation among the immunoprecipitation, electrophoretic, and chemical inhibition methods was good (r greater than 0.991), although the differences between means were statistically significant (except for the comparison of the two chemical inhibition methods). Because of its ease, low cost, and precision, we recommend the perchlorate method for routine use.

Does serial determination of lactate dehydrogenase isoenzymes 1 and 2 ratios contribute to the diagnosis of acute myocardial infarction?

Journal Article Does serial determination of lactate dehydrogenase isoenzymes 1 and 2 ratios contribute to the diagnosis of acute myocardial infarction? Get access Z Rotenberg, Z Rotenberg Israel and Ione Massada Center for Heart Diseases, Dept. of Med. A, Petah Tikva, Israel Search for other works by this author on: Oxford Academic Google Scholar I Weinberger, I Weinberger Israel and Ione Massada Center for Heart Diseases, Dept. of Med. A, Petah Tikva, Israel Search for other works by this author on: Oxford Academic Google Scholar E Davidson, E Davidson Israel and Ione Massada Center for Heart Diseases, Dept. of Med. A, Petah Tikva, Israel Search for other works by this author on: Oxford Academic Google Scholar J Fuchs, J Fuchs Israel and Ione Massada Center for Heart Diseases, Dept. of Med. A, Petah Tikva, Israel Search for other works by this author on: Oxford Academic Google Scholar O Sperling, O Sperling Israel and Ione Massada Center for Heart Diseases, Dept. of Med. A, Petah Tikva, Israel Search for other works by this author on: Oxford Academic Google Scholar J Agmon J Agmon Israel and Ione Massada Center for Heart Diseases, Dept. of Med. A, Petah Tikva, Israel Search for other works by this author on: Oxford Academic Google Scholar Clinical Chemistry, Volume 34, Issue 7, 1 July 1988, Pages 1506–1507, https://doi.org/10.1093/clinchem/34.7.1506a Published: 01 July 1988

Selective determination of lactate dehydrogenase isoenzyme 1 in serum after inhibition by 1,6-hexanediol.

A rapid selective method for measuring the activity of lactate dehydrogenase isoenzyme LD-1 in serum by using 1,6-hexanediol as an inhibitor of the M-subunit was developed. Hexanediol was added to serum at a final concentration of 0.7 mol/l. After incubation at 30 degrees C for 15 min, the activity was measured with an automatic analyser. The inter-assay coefficient of variation was 6.9% for the lactate dehydrogenase isoenzyme LD-1 measurement. The results obtained from the sera of 100 patients analysed by the proposed selective method and by the conventional electrophoretic method, respectively, showed an excellent correlation. This selective method was used to determine the lactate dehydrogenase isoenzyme LD-1 activity of sera from patients with acute myocardial infarction, and the results were correlated well with those obtained by the immunological, Roch Isomune method. Addition of 1,6-hexanediol did not affect the measurement of activities of other enzymes such as alkaline phosphatase, gamma-glutamyltransferase, aspartate aminotransferase and alanine aminotransferase.

Determination of Lactate Dehydrogenase Isoenzyme (LD-1) Using Flow Injection Analysis with Electrochemical Detection after Immunochemical Separation

Abstract A flow system has been developed for a fast and convenient assay of LD-1 and total LD in serum. The LD-1 is separated immunochemically and subsequently determined amperometrically by a thin layer transducer in a flow system. Potassium ferricyanide was used as a mediator at a potential of 0.07 V vs Ag/AgCl to eliminate any protein fouling of the electrode. A kinetic measurement helps to diminish several interferences in the electrode response. Thus, it is possible to determine the LD-1 or LD-1/total LD conveniently and accurately even at low activities.

Lactate Dehydrogenase Isoenzyme 1

• A rapid immunochemical method (Isomune-LD) for the determination of lactate dehydrogenase isoenzyme 1 (LD₁) activity in serum was investigated, and the sensitivity and specificity of the assay were determined for the diagnosis of a myocardial infarction (Ml). Multiple serum samples were obtained (on admission and at 12, 24, and 48 hours) from 85 patients (30 with proven Ml and 55 without Ml). The sensitivity of LD₁on admission and at 12, 24, and 48 hours was 43%, 84%, 96%, and 100%, respectively, and the specificity at these same time intervals was 95%, 100%, 97%, and 97%, respectively. Thus, the determination of LD₁by this rapid immunochemical method appears to allow a rapid laboratory evaluation of Ml with a high degree of sensitivity and specificity. (<i>Arch Intern Med</i>1983;143:434-436)

Lactate dehydrogenase isoenzyme 1. A rapid assay for ischemic myocardial disease

A rapid immunochemical method (Isomune-LD) for the determination of lactate dehydrogenase isoenzyme 1 (LD1) activity in serum was investigated, and the sensitivity and specificity of the assay were determined for the diagnosis of a myocardial infarction (MI). Multiple serum samples were obtained (on admission and at 12, 24, and 48 hours) from 85 patients (30 with proven MI and 55 without MI). The sensitivity of LD1 on admission and at 12, 24, and 48 hours was 43%, 84%, 96%, and 100%, respectively, and the specificity at these same time intervals was 95%, 100%, 97%, and 97%, respectively. Thus, the determination of LD1 by this rapid immunochemical method appears to allow a rapid laboratory evaluation of MI with a high degree of sensitivity and specificity.

Immunochemical and electrophoretic methods for determination of lactate dehydrogenase isoenzyme 1 compared.

Journal Article Immunochemical and electrophoretic methods for determination of lactate dehydrogenase isoenzyme 1 compared. Get access S S Fike S S Fike Search for other works by this author on: Oxford Academic Google Scholar Clinical Chemistry, Volume 26, Issue 6, 1 May 1980, Pages 794–795, https://doi.org/10.1093/clinchem/26.6.794a Published: 01 May 1980

Immunochemical and electrophoretic methods for determination of lactate dehydrogenase isoenzyme 1 compared.

Immunochemical and electrophoretic methods for determination of lactate dehydrogenase isoenzyme 1 compared.

Spurious brain creatine kinase in serum from patients with renal disease.

Creatine kinase isoenzyme I(BB) is generally not detectable in normal serum, and its occurrence in serum has been documented in only a few disease states. In particular, increased activity of this isoenzyme has been reported in association with chronic renal failure, hemodialysis, and renal transplantation. The present study demonstrates that the apparent creatine kinase observed in the serum of such renal patients is an artifact, observed as a result of measuring creatine kinase isoenzymes by fluorescence. Our observations resemble those of McKenzie et al. [Clin. Chim. Acta 70, 333(1976)] concerning an artifact in the fluorometric determination of lactate dehydrogenase isoenzymes in the sera of patients with end-stage renal failure. The artifact binds to albumin, is not a protein, and occurs in some normal sera at very low concentrations. This artifact can be mistakenly identified as isoenzyme I in renal-disease patients if CK isoenzymes are determined fluorometrically.

Evaluation of lactate dehydrogenase isoenzyme patterns in serum of patients undergoing cardiac surgery

Serial determinations of serum lactate dehydrogenase (LD) isoenzymes were performed in 50 patients undergoing cardiac surgery for coronary artery bypass and heart valve replacement. A sequence of LD isoenzyme patterns was established in the patients with uncomplicated recovery. These patterns served as controls for the detection of abnormal patterns associated with clinical complications. Perioperative myocardial infarction was detected in seven patients by a characteristic reversal of the LD1:LD2 ratio. These studies established that accurate determination of LD isoenzymes improved their diagnostic specificity. These isoenzymes were determined by (1) using the same technique for tissues and for serum samples, (2) applying a volume of serum containing a standard amount of enzymatic activity, and (3) making a clear separation, which allows accurate quantitation. This test can provide useful information to the surgeon for the evaluation of operative procedures and to the clinician for the appropriate management of the patient undergoing cardiac surgery.

Automated kinetic determination of lactate dehydrogenase isoenzymes in serum.

A steady-state kinetic method has been revised for measuring lactate dehydrogenase isoenzyme activities, which relates the inhibition of heart-type isoenzyme activity to the overall isoenzyme composition of the enzyme subunits. The method depends on the pH-dependent formation of an inhibitory ternary complex by the heart-type isoenzyme with NAD+ and pyruvate (if the reaction is measured by NADH oxidation). A preincubation step in the previous method is eliminated. The isoenzymes are measured by measuring the reduction of pyruvate in two different concentrations, which favor either the total or fractional activity, depending on the concentrations of pyruvate and the percentage of heart-type subunits. The method has been adapted to a centrifugal analyzer, which has speeded automated isoenzyme determinations, with an accuracy comparable to that for electrophoretic methods.

H. A. Zondag. Determination and diagnostic significance of lactate dehydrogenase isoenzymes. 118 p., 24 fig. Assen, 1968

This book is the first successful attempt to summarize the literature data and the author's own research on the definition and diagnostic evaluation of lactate dehydrogenase isoenzymes. The monograph is written on the basis of the analysis of 4000 studies and accounting of the work of exclusively foreign laboratories and clinics. Among the 290 cited works in the bibliography there is no reference to any work of Soviet scientists. Meanwhile, in our country, the biochemical determination of lactate dehydrogenase isoenzymes is successfully used, especially in the pathology of the cardiovascular system.

The determination of lactate dehydrogenase isoenzymes in normal human muscle and other tissues

1. A technique has been developed, based on preferential inhibition by urea, for determining the amounts and proportions of the M and H sub-units of lactate dehydrogenase (referred to as LDH-M and LDH-H respectively) in human tissues, including muscle. 2. There was good agreement between the results obtained with urea inhibition and those obtained with starch-gel electrophoresis. 3. With increasing age there was a significant decrease in the total amount of lactate dehydrogenase and the amount of LDH-M in skeletal muscle. This could not be accounted for by the replacement of functioning muscle tissue by fibrous connective tissue. 4. The proportion of LDH-M was less in certain muscles (e.g. soleus and extra-ocular) than in other muscles (e.g. gastrocnemius and rectus abdominis). 5. The proportions of LDH-M and LDH-H did not differ significantly in different superficial limb muscles and were not significantly affected by either age or sex. 6. Specimens of muscle from 86 different individuals (all Europeans) have been subjected to electrophoresis, but no variants of lactate dehydrogenase isoenzymes have been found.

The determination of lactate dehydrogenase iso-enzymes using the microzone cellulose acetate system

o 1. A rapid and simple technique using the Beckman Microzone celluloseacetate system for the electrophoretic separation of serum lactate dehydrogenase isoenzymes is described. 2. The isoenzymes are determined using the coupled nitro-blue-tetrazoliumreaction followed by densitometry of the formazan bands. Up to eight sera can be analysed for lactate dehydrogenase in one hour. The method is compared with that of starch gel electrophoresis. 3. Isoenzyme studies of normal individuals and patients are reported.