Two methods compared for measuring LD-1/total LD activity in serum

We present evidence for the utility of an improved assay for the activity of lactate dehydrogenase (EC 1.1.1.27) isoenzymes 1 and 2 in serum, involving inhibition of the H-subunit of LD by pyruvate at pH 7.1. Results correlate well with the LD-1/total LD ratio as evaluated by immunological assay and, as an index to infarct, the method is superior to either the change in CK-MB activity or to the LD-1 activity or to a combination of these tests, as is the percentage of LD-1 to total LD activity. Moreover, the percentage inhibition of LD activity by pyruvate may have an advantage over other methods of isoenzyme fractionation because of its smaller population CV for patients with acute myocardial infarction than is true of other methods. We also demonstrate how, using a linear discriminant analysis, we compared this method with alternative methods. We determined that evaluation of CK-MB isoenzyme contributes no information in addition to that obtained from the LD-1 isoenzyme.     

Lactate dehydrogenase isoenzyme-1/total ratio: accurate for determining the existence of myocardial infarction

We have gradually revised our medical protocols for measuring creatine kinase MB isoenzyme (CK-MB) and lactate dehydrogenase isoenzyme-1 (LD-1) because of identifiable problems in the use of an interpretation of CK-MB isoenzyme associated with slowly evolving or small myocardial infarct, the use of thrombolytic therapy, or burn and trauma, each of which affects the rate of appearance and composition of isoenzymes present. Despite recent evidence of the efficacy of LD-1 isoenzyme measurement in the first 12 to 24 h of myocardial infarction, this test is not widely used because of overstated assumptions about the value of CK-MB. Here we studied the adequacy of the current isoenzyme assays by determining the value of CK-MB and LD-1 at optimum serum sampling times and establishing the contribution of individual and combined predictors to diagnostic efficiency. We conclude that the LD-1/total LD activity ratio in serum is superior to measurement of CK-MB or LD-1, or both, in the diagnosis of acute myocardial infarction. Moreover, this ratio is most valuable when interpretation of the result for CK-MB isoenzyme is equivocal in patients with small or evolving myocardial infarcts.     

Changes in the ratio of lactate dehydrogenase isoenzymes 1 and 2 during the first day after acute myocardial infarction

It is known that the ratio of isoenzyme 1 to total lactate dehydrogenase (LD, EC 1.1.1.27) in serum is increased in all patients with acute myocardial infarction within 24 h of the infarct. We now show that the LD-1/LD-2 ratio for serum more promptly indicates acute myocardial infarction, being for most patients equivalent to measurement of creatine kinase (EC 2.7.3.2) isoenzyme 2 (CK-2, CK-MB) in serum. Of 128 patients with a confirmed diagnosis of myocardial infarction, 66 had normal values for all "cardiac" enzymes at the time of admission, but greater than 75% of them showed a parallel increase in values for CK-2 and the LD-1/LD-2 ratio. Of the 26 patients who had one or more abnormal values for cardiac enzymes on admission, 95% showed a parallel increase in CK-2 and the LD-1/LD-2 ratio, the median time for the beginning of these changes being 9 h from the onset of chest pain. The remaining 36 patients were excluded from the study because CK-2 decreased after admission or because the time of onset of chest pain was uncertain.     

"Flipped" patterns of lactate dehydrogenase isoenzymes in serum of elite college basketball players

We kinetically measured total lactate dehydrogenase (LD, EC 1.1.1.27), total creatine kinase (CK, EC 2.7.3.2), and aspartate aminotransferase (AST, EC 2.6.1.1.) in 16 elite college basketball players, before the competition season and not in close temporal relation to near-maximal exercise, and in 17 healthy non-athlete controls. LD isoenzymes were determined by both electrophoretic and immunoprecipitation methods. CK-MB isoenzyme was measured electrophoretically. We found significantly higher mean LD-1 values and LD-1/LD-2 ratios in the players than the controls: 31.6 (SD 3.7)% vs 25.8 (SD 3.2)% (P less than 0.005) and 1.1 (SD 0.13) vs 0.87 (SD 0.16) (P less than 0.001), respectively. A "flipped" LD pattern (LD-1 greater than LD-2) was found in half the players and in six of the eight black athletes, but in only two of the control group and in none of the black controls. Mean CK activity in serum exceeded normal values in the serum of the athletes and was higher in comparison with the control group [274 (SD 156) vs 103 (SD 82) U/L]. Mean CK was significantly higher in the eight athletes with the flipped LD pattern than in those with LD-1 less than LD-2 [322 (SD 163) vs 180 (SD 98) U/L; P = 0.05], and also in comparison with CK in the two controls with flipped LD pattern. We saw no significant difference in mean CK between the nine players with normal immunochemical LD-1/LD ratios and the seven players with above-normal ratios. CK-MB was not detected in either athletes or controls. None of the players had any clinical or electrocardiographic evidence for myocardial ischemia or infarction. Evidently the flipped LD pattern usually found in patients with acute myocardial infarction and reported in some athletes after extreme exercise such as ultra-marathon running may also be found in athletes who are in their "basal fitness shape" but who are not involved in competitive physical activity.     

Increased activity of creatine kinase isoenzyme MB in a theophylline-intoxicated patient

A 78-year-old woman had increased activities of creatine kinase (CK; EC 2.7.3.2) and CK-MB isoenzyme in her serum, associated with severe theophylline intoxication. The time course for CK-MB activity was similar to that from an acute myocardial infarction. Clinical findings, however, including electrocardiograms, did not support the diagnosis of myocardial infarction. We suggest caution in interpreting CK-MB results in severe theophylline intoxication.     

Increased lactate dehydrogenase isoenzyme 1 in serum and tumor tissue of a patient with small-cell carcinoma

Histological examination of supraclavicular lymph node tissue obtained at biopsy from a 63-year-old man disclosed metastatic small-cell carcinoma. On admission and for four days subsequently, total lactate dehydrogenase (LD; EC 1.1.1.27) activity in serum was 6.5 times normal; studies of LD isoenzyme showed persistently increased LD-1, with LD-1 greater than LD-2. Isoenzyme electrophoresis of tissue homogenates prepared from the patient's tumor also showed the LD-1 greater than LD-2 pattern. Isoenzyme studies for supraclavicular lymph node tissue from five control subjects showed contrasting isoenzyme patterns as compared with the patients in whom LD-2, LD-3, and LD-4 predominated. Because these abnormalities were persistent, they differ from the temporal sequence for LD usually seen in myocardial infarction. This emphasizes the importance of repetitive sampling for clinical interpretation of data on this enzyme.     

Atypical patterns of lactate dehydrogenase isoenzymes in acute myocardial infarction

Total lactate dehydrogenase (LD; EC 1.1.1.27) activity in serum and LD isoenzymes were quantified in 190 patients with acute myocardial infarction (AMI) 24, 48, and 72 h after admission. In 90% of the 570 blood specimens an LD isoenzyme pattern typical of AMI (LD-1/LD-2 greater than 0.76) was found. The other 56 blood specimens showed an LD isoenzyme pattern atypical of AMI (LD-1/LD-2 less than 0.76). They were divided into three groups: 28 specimens with isomorphic pattern (relative increase in all five LD isoenzymes); 18 with relatively increased LD-3 proportion (greater than 35%); and 10 specimens with increased LD-5 proportion (greater than 10%). No difference was found in mean total LD activity in serum between the typical isoenzyme group and the three atypical groups. The LD isomorphic pattern was found in 60% of AMI patients complicated by cardiogenic shock. Fifty percent of AMI patients admitted with pulmonary edema showed increased LD-3 proportion and half of the patients with AMI and congestive heart failure, predominant right, demonstrated increased LD-5 proportion. We conclude that although most patients with AMI present at diagnosis with a typical LD isoenzyme pattern, it is important to recognize that some may present with atypical LD isoenzyme patterns, which may be associated with specific AMI complications.     

Determination, by radioimmunoassay, of the mass of lactate dehydrogenase isoenzyme 1 in human serum and of its rate of removal from serum after a myocardial infarction

In this radioimmunoassay of lactate dehydrogenase-1 (LD-1; EC 1.1.1.27) in human serum we use a commercial LD-1-selective assay system and a goat antiserum. We have determined the fractional rate of disappearance from serum and the half-life of LD-1, in terms of both enzyme activity and enzyme mass, in 21 myocardial infarction patients. Our evidence suggests that this isoenzyme is inactivated in serum. Furthermore, our data suggest that the conventionally accepted half-life of about 110 h for serum LD-1 activity may grossly overestimate the actual LD-1 half-life in many post-myocardial infarction patients.     

Lactate dehydrogenase isoenzyme 1: Its usefulness as a screening test in diagnosis of myocardial infarction

Immunological assay of LD-1 activity provides a quantitative measurement of the type of lactate dehydrogenase (LD, EC 1.1.1.27) activity characteristic of myocardial origin. Using this test, a laboratory diagnosis of myocardial infarction can be either ruled out or confirmed in approximately 75% of patients in whom this diagnosis is suspected, without electrophoretic separation of creatine kinase (CK, EC 2.7.3.2.) and LD isoenzymes. Normal total CK and LD activities cannot be used to rule out myocardial infarction since CK-MB and LD-1 may have increased although total activities remain within their reference ranges. LD-1 activity increases as quickly as CK-MB following the onset of pain in the majority of patients but it remains elevated longer giving a greater period of time during which the diagnosis of myocardial infarction can be confirmed.     

Use of purified lyophilized human lactate dehydrogenase isoenzymes in a study of the measurement of lactate dehydrogenase activity

We examined the stability of human lactate dehydrogenase (EC 1.1.1.27; LD) isoenzymes 1, 2, and 3--purified to specific activities of about 200 kU/g--when lyophilized in a buffered stabilized matrix of bovine albumin. Each isoenzyme was prepared at two activity concentrations and stored at -20, 4, 20, 37, and 56 degrees C for as long as six months. LD-1 activity decayed with zero-order kinetics, LD-2 and LD-3 with first-order kinetics. The extrapolated half-lives of these preparations at -20 degrees C varied between 80 and 530 years. Stability of reconstituted samples stored at 4 degrees C was excellent for LD-1 but poor for LD-2 and LD-3. We suggest that preparations of human LD-1 be further investigated as a possible reference material.     

Increase in lactate dehydrogenase isoenzyme-1 in plasma in pediatric malignancy

We investigated 28 cases of pediatric malignancy in which total lactate dehydrogenase (LD, EC 1.1.1.27) activities were increased and isoenzyme LD-1 exceeded LD-2 (flipped pattern). Of these, 11 had a flipped pattern at presentation and 17 showed a flipped pattern during chemotherapy. Those with flipped patterns at presentation were four with germ-cell tumors, one with acute lymphocytic leukemia, and six with nephroblastomas (Wilms tumor). Four of five nephroblastoma homogenates contained predominantly LD-1; one revealed a structurally normal LD-1 with normal kinetics. We conclude that an increase in LD with a flipped pattern is common in nephroblastoma and, in addition, may develop in cancer patients treated with chemotherapy.     

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.     

Unexplained increase in serum creatine kinase isoenzyme MB activity in a lung cancer patient

In this case of mixed small cell--large cell cancer of the lung in an elderly woman, creatine kinase (EC 2.7.3.2) isoenzymes were assayed serially because of chest pain. The proportions of serum CK-BB and CK-MB isoenzyme activities were persistently above normal (CK-MB 10-18%, normal less than 5%). Electrocardiograms revealed no signs of ischemia or infarction. At autopsy no gross or microscopic infarction or inflammation of the heart was seen. There was also no infarction of smooth or skeletal muscle. The tumor was the probable source of most of the circulating CK-MB isoenzyme. Future cases may pose a similar diagnostic dilemma: differentiating creatine kinase that is present as a result of myocardial infarction from tumor-related CK-MB. Whether or not CK-MB assay could be useful in detecting tumors remains to be investigated.     

Stability of lactate dehydrogenase at different storage temperatures

The effect of storing human serum, cord blood serum or heparinized plasma at 25 degrees C, 4 degrees C & -20 degrees C on the activity and isoenzyme distribution of lactate dehydrogenase (LD) was studied. Cellulose acetate and agarose electrophoresis, as well as an immunochemical inhibition technique, were used for isoenzyme quantification. In contrast to previous reports, cryo-instability was found only in specimens stored at 4 degrees C. Serum specimens stored at 25 degrees C and -20 degrees C retained 74% and 87% of total activity after 45 days of storage. LD-1 was stable at all three temperatures, with a maximum loss of 10%. LD-2, LD-3, LD-4, & LD-5 were most labile at 4 degrees C. Specimens that are to be analyzed for total LD or LD isoenzymes should be stored frozen or, if necessary, at room temperature, but not in a refrigerator. Thus, separate storage of specimens for cardiac isoenzymes (LD & creatine kinase) is not necessary. This may eliminate a possible source of falsely elevated LD-1/LD-2 ratios, as well as reducing the labor factor and the corresponding cost of cardiac isoenzyme determinations.     

Increased serum lactate dehydrogenase isoenzyme 1 and "flipped" LD-1/LD-2 ratio in myopathy associated with partial carnitine palmitoyltransferase deficiency

We describe a case of a limb-girdle myopathy presenting with myoglobinuria. A partial deficiency of muscle carnitine palmitoyltransferase (EC 2.3.1.21) may also have been present. All "muscle-type" serum enzymes were markedly increased (to between 30- and 400-fold their respective upper reference limits) and creatine kinase (EC 2.7.3.2) isoenzyme 2 (CK-MB) was increased 130-fold but was still less than 2% of the total creatine kinase activity. The isoenzyme pattern of lactate dehydrogenase (EC 1.1.1.27) in serum was "anodic," with isoenzyme 1 greater than isoenzyme 2--an unusual pattern for myopathies. The possible physiological basis for such a finding is discussed.     

Creatine kinase isoenzymes of mitochondrial origin in human serum.

We measured creatine kinase (EC 2.7.3.2) activity in 1009 serum samples from 538 patients in the intensive-care units of the University of Texas Medical Branch hospitals. Creatine kinase isoenzymes migrating cathodal to skeletal muscle creatine kinase (CK-MM) on cellulose acetate electrophoresis were found in sera from 14 of the 538 patients. Creatine kinase, lactate dehydrogenase (EC 1.1.1.27), aspartate aminotransferase (EC 2.6.1.1), and alanine aminotransferase (EC 2.6.1.2) activities were abnormally increased in these 14 patients. Liver lactate dehydrogenase isoenzyme (LDH5) and cardiac creatine kinase isoenzyme (CK-MB) were abnormally increased in 12 and eight of these patients, respectively. Ten of the 14 patients died during their hospital admission. We believe the creatine kinase isoenzymes that migrated cathodal to skeletal muscle creatine kinase (CK-MM) were of mitochondrial origin.     

Inhibition of lactate dehydrogenase isoenzymes by sodium perchlorate evaluated

We evaluated a method of measuring lactate dehydrogenase isoenzyme 1 (LD-1) selectively (Clin Chem 1987;33:991-2), in which all other LD isoenzymes were inhibited by adding sodium perchlorate to the reaction medium to a final concentration of 0.825 mol/L. In this study we used the different isoenzymes purified from human autopsy tissue and found that the originally published amount of inhibitor (a) increased the original LD-1 activity and (b) did not eliminate all enzyme activity of LD-2 and LD-3. Interference by LD-2 was further demonstrated. Thus we conclude that this method cannot be used for the selective determination of LD-1 because its results do not accurately reflect the original LD-1 activity.     

Changes in serum CK-MB mass after coronary artery bypass surgery

We assessed the release of creatine kinase MB as both mass and activity during the postoperative period following cardiac surgery. CK-MB mass was determined by enzyme immunoassay using reagents obtained from Hybritech. CK-MB activity was determined both by agarose electrophoresis and by an immunochemical method. Fifty-five patients who underwent coronary artery bypass surgery and 52 control subjects who had orthopedic surgery were selected for study. Serial serum samples were collected following surgery and total LD, CK, AST, LD-1, CK-MB mass, and CK-MB activity determined. Results were compared to each other and to surgical parameters. All patients exhibited significant CK-MB mass and activity after surgery and peak serum levels were 6-94 micrograms/L and 12-84 U/L, respectively. CK-MB mass correlated with CK-MB activity on paired samples (r = 0.94). Total AST and CK activities correlated with CK-MB mass (r = 0.60, and 0.63, respectively). Peak levels of CK-MB mass correlated significantly with peak MB activity (r = 0.88), peak LD-1 (r = 0.62), peak AST (r = 0.71), and time on pump (r = 0.54). Similar correlations were also seen between peak CK-MB activity and these parameters. No relationship could be identified between extent of CK-MB mass release and number of grafts, degree of hypothermia, or minimum PaO2. The time course of CK-MB mass release exhibited 85% concordance with CK-MB activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Measuring creatine kinase MB isoenzyme in a maintenance hemodialysis population: chemiluminometric immunoassay and electrophoresis compared

In an effort to clarify the issue of potentially false increases in creatine kinase (EC 2.7.3.2) MB isoenzyme (CK-MB) in uremia, we evaluated the CK profile of 84 persons undergoing chronic maintenance hemodialysis. We compared the performance of a new commercial two-site chemiluminometric immunoassay of CK-MB (Magic Lite; Ciba Corning Diagnostics) with that of electrophoresis on agarose gel (Cardio Trak-CK; Corning Medical). Results of the new chemiluminometric immunoassay for samples from hemodialysis patients correlated well with those of the electrophoretic method (r = 0.86, P less than 0.001), showing that neither substances in the serum of uremic patients nor CK-MM isoenzyme give false-positive increases in CK-MB isoenzyme. Our evidence suggests that the chemiluminometric method may be more specific than is electrophoresis in establishing absolute CK-MB values in the diagnosis of suspected myocardial injury in this population.     

Clinical effectiveness of the Du Pont aca measurement of creatine kinase MB in serum from patients in a coronary-care unit

We evaluated the clinical effectiveness of measuring creatine kinase (CK; EC 2.7.3.2) isoenzyme MB and lactate dehydrogenase (LD; EC 1.1.1.27) isoenzymes in diagnosis of acute myocardial infarction. We used an agarose electrophoresis method to measure CK and LD isoenzymes and the Du Pont aca column method to measure CK-MB. Serial blood specimens were drawn from 100 patients consecutively admitted to our Coronary Care Unit. Because of the low diagnostic specificity for CK-MB measurements by both agarose electrophoresis and the discrete-analysis method, as compared with reported values, we re-evaluated our isoenzyme data by using Receiver Operating Characteristic curves. Such analysis of the data established optimal decision levels of greater than or equal to 25 U/L and greater than or equal to 18 U/L plus greater than or equal to 6% of total CK for serum CK-MB measured by the agarose electrophoresis and the aca methods, respectively, and an optimal decision level of greater than or equal to 0.92 for the ratio of LD 1/2 measured after agarose electrophoresis. At these decision levels we obtained a sensitivity of 100%, 100%, and 95% and a specificity of 94%, 92%, and 90% for CK-MB (agarose electrophoresis), CK-MB (aca), and the LD 1/2 ratio, respectively.