Interpretation of changes in the activity of creatine kinase MB isoenzyme in serum after coronary artery bypass grafting

Measurement of CK-MB in the sera of patients undergoing coronary artery bypass grafting shows a significant increase above pre-operative levels in all cases. However, the timing of the peak post-operative activity and its level allow criteria to be proposed which differentiate between two classes of patients: those with myocardial infarction or lesser myocardial damage, in whom peak activities are seen in specimens taken 21 hours after operation; and those in whom peak activities are found in specimens taken 4-7 hours after operation, presumably reflecting reversible myocardial changes. Activity peaks of more than 50 U/l occurring at 21 hours are considered to support a diagnosis of myocardial infarction.     

Mass concentration of creatine kinase MB isoenzyme and lactate dehydrogenase isoenzyme 1 in diagnosis of perioperative myocardial infarction after coronary bypass surgery

Recent advances in methodology allow the mass concentration of creatine kinase MB isoenzyme (CK-MB), and of lactate dehydrogenase isoenzyme 1 (LD1) to be determined quickly and easily as routine, emergency tests. We evaluated these tests as diagnostic criteria of perioperative myocardial infarction (PMI) after coronary bypass surgery. These tests were compared with the usual measurements of CK-MB activity by immunoinhibition and LD1 by electrophoresis and with other biological markers of myocardial infarction such as total CK, total LD, and aspartate aminotransferase. Sixty-one patients who underwent coronary bypass grafting were followed pre- and postoperatively by enzyme determinations and electrocardiography; a subgroup was monitored by myocardial scintigraphy. CK-MB mass appeared to be the best marker of PMI during the first 48 h, although LD1 was the marker of choice from days 2 to 4.     

Assay of creatine kinase isoenzyme MB in serum with time-resolved immunofluorometry

We describe the first time-resolved immunofluorometric assay for creatine kinase (EC 2.7.3.2) isoenzyme MB (CK-MB) in serum. The assay is based on the formation of the complex: solid-phase anti-CK-MB-CK-MB-biotinylated anti-CK-BB-streptavidin-BCPDA-Eu3+, where anti-CK-MB and anti-CK-BB are monoclonal antibodies against the CK isoenzymes MB and BB, respectively, and BCPDA is the europium chelator 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid. The solid-phase complex is fluorescent and is measured on the dry solid-phase (microtiter well) in a specially designed time-resolved fluorometer that uses laser excitation. The assay requires 25 microL of serum and is not affected by the presence of either CK-MM (up to 5000 micrograms/L) or CK-BB (up to 1000 micrograms/L) in the sample. Precision and accuracy indices for the assay were satisfactory.     

Proteinase K inactivation of cytosolic aspartate aminotransferase isoenzyme for measurement of human serum mitochondrial aspartate aminotransferase

We studied a new proteinase K assay method for human serum mitochondrial aspartate aminotransferase. We found that proteinase K showed no inactivation of human mitochondrial aspartate aminotransferase isoenzyme and complete inactivation of cytosolic aspartate aminotransferase. Previous studies have shown that selective proteolytic measurement for mitochondrial aspartate aminotransferase in serum using the protease 401 cleaved peptide bond at Leu 20 from the amino-terminal bond shows complete inactivation of cytosolic aspartate aminotransferase and slight inactivation of mitochondrial aspartate aminotransferase isoenzyme, depending on protease concentration. In this investigation, we found that the proteinase K method does not depend on protease concentration. The proteinase K enzyme inactivation of cytosolic aspartate aminotransferase is caused by the cleavage of the peptide bond at lieu 21 from the aminoterminal bond. In studies with various animal cytosolic aspartate aminotransferase isoenzymes, proteinase K almost completely inactivated cytosolic aspartate aminotransferase. Precision and correlation using proteinase K for measurement of serum mitochondrial aspartate aminotransferase in human showed a good coefficient of variation (within-run <4.45%) and a coefficient of correlation of r = 0.985 (N = 125). © 1993 Wiley-Liss, Inc.     

Relation of creatine kinase isoenzyme MB to postoperative electrocardiographic diagnosis in patients undergoing coronary-artery bypass surgery.

We compared (a) the frequency of detection of isoenzyme MB of creatine kinase (CK; EC 2.7.3.2) in serum of patients undergoing coronary-artery bypass surgery, (b) the interval during uhich its activity was supranormal in serum, and (c) an index of the amount of CK released into blood ("CK-MB area") with postoperative electrocardiographic changes in 80 patients. The frequency of detection of CK-MB is a function of frequency of sampling during the early postoperative period. Because the duration of appearance and the calculated CK-MB area increased as the electrocardiogram became more specific for infarction (p less than 0.01), a twice-daily sampling schedule proved clinically relevant. Only 5.4% of patients had electrocardiographic evidence of infarction when CK-MB was absent by the second postoperative morning. When CK-MB was still detected at that time, 69.6% of patients had persistent new Q waves, consistent with infarction. In three patients who died postoperatively, significant myocardial necrosis was demonstrated. All three had had persistently increased values for CK-MB, related to electrocardiographic changes of infarction in one patient and ischemic changes in two. Evidently CK-MB is a more sensitive indicator of myocardial necrosis than the electrocardiogram and CK-MB area should be a useful criterion in evaluating methods of intra-operative myocardial protection.     

Immunochemical extraction and automated measurement of plasma creatine kinase MB isoenzyme and creatine kinase MB2 isoform

Measurement of creatine kinase MB (CK-MB) and its isoforms CK-MB₂ and CK-MB₁ are now applied in the diagnosis of acute myocardial infarction (AMI). The most common approach for analysis includes RIA, IRMA, and electrophoresis, all of which may be time-consuming. This study examines determination of CK-MB and CK-MB₂ by a rapid immunochemical extraction method followed by an automated measurement for both analytes. The automated method was sensitive to 2 U/L, linear to 180 U/L, and gave excellent interassay precision (<10% CV). Interference studies indicated that bilirubin, hemolysis, and lipemia caused analytical problems as did the presence of high activities of other CK isoenzymes, notably CK-MM and CK-BB, requiring dilution of samples prior to analysis. Application of immunochemical extraction gave a reference interval of CK-MB (0–2.5 U/L) and CK-MB₂ (0.1–1.4 U/L) for blood donors (20–60 years), peak levels for ruled-out AMI patients of CK-MB (0.5–7.3 U/L) and CK-MB₂ (0.3–4.9), peak levels for ruled-in AMI patients of CK-MB (80–174 U/L) and CK-MB₂ (80–155 U/L). Coronary artery bypass patients (n = 24) and all trauma patients (n = 14) also demonstrated elevations in CK-MB and CK-MB₂, whereas only five of the trauma patients demonstrated increased CK-MB by IRMA. In patients (n = 7) having increased total CK and normal CK-MB by IRMA, the extraction assay for CK-MB and CK-MB₂ yielded increased values in all patients. This new approach to CK-MB and CK-MB₂ analysis can be performed within 30 minutes of sample receipt. J. Clin. Lab. Anal. 11:163–168, 1997. © 1997 Wiley-Liss. Inc.     

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.     

Determination of mitochondrial aspartate aminotransferase in serum

Two specific and sensitive immunoassay methods for the determination of mitochondrial aspartate aminotransferase (m-AST) are described. One is a sandwich enzyme immunoassay which measures immunologically active m-AST using polystyrene balls coated with anti-m-AST antibody and peroxidase-labelled anti-m-AST antibody as the second antibody. The detection limit of this assay was 10 micrograms/l. The other is a paper disk method which measures catalytically active enzyme bound to anti m-AST antibody-conjugate paper disks. The calibration curve was linear up to 250 U/l. These assay methods were used to monitor the level of m-AST in serum. From measurements obtained by both methods, the correlation between the concentration of m-AST protein and its activity was poor (liver diseases, r = 0.539; myocardial infarction, r = 0.774) confirming that an inactive form of m-AST exists in serum, and that the specific activity of serum m-AST differs in individual diseases.     

Serum creatine kinase isoenzyme MB concentration after endomyocardial biopsy

Serum total creatine kinase (CK), CK-MB and myoglobin (Mb) were serially determined in 17 patients who underwent endomyocardial biopsy. Mean total CK levels increased from 36 +/- 27 U/l 30 min before biopsy to a maximum of 112 +/- 77 U/l 8 h following the procedure (p less than 0.05). Similarly, Mb concentrations rose from 57 +/- 55 micrograms/l to 119 +/- 57 micrograms/l 30 min after biopsy (p less than 0.05). Normalization of total CK and Mb levels occurred within 16 and 8 h, respectively. A new immunoenzymetric assay (IEMA) was used to measure the mass concentration of the CK-MB molecule. The initial CK-MB levels were 0.2 +/- 0.4 microgram/l; a small but significant elevation was recorded as early as 2 h after biopsy (1.6 +/- 1.5 micrograms/l, p less than 0.05). CK-MB returned to initial concentration 16 h after the beginning of the procedure. Comparison with the maximum CK-MB levels recorded in 16 myocardial infarction patients (258 +/- 172 micrograms/l, range 90-680 micrograms/l) indicated that the modest increase of CK-MB level detected after biopsy probably reflects a limited endomyocardium lesion at the sampling site, excluding any significant myocardial damage. Total CK and Mb, which showed more pronounced elevations than CK-MB, are likely to originate from other sources than the myocardium.     

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.     

Comparison of 3 methods of analysis of the MB isoenzyme of creatine kinase in serum.

A new batch method for quantitating the MB isoenzyme of creatine kinase in human serum using DEAE Sephadex is described and compared with 2 electrophoretic methods using (1) polyacrylamide gel and (2) agarose film. Their relative merits for use as a routine method in clinical laboratories are discussed.     

Immunochemiluminometric assay of creatine kinase MB with a monoclonal antibody to the MB isoenzyme

Previous two-site immunometric assays for creatine kinase (CK; EC 2.7.3.2) MB isoenzyme have been based on formation of a "sandwich" complex involving CK-MB and antibodies that recognize the CK-MM and the CK-BB isoenzymes. Single-incubation model assays of CK-MB with these antibodies were susceptible to interferences by CK-MM and CK-BB. We produced two anti-CK-MB monoclonal antibodies and studied their suitability for two-site assays. Both antibodies were compatible with anti-CK-MM and anti-CK-BB, but not with each other. Using anti-CK-MB as the tracer antibody eliminated the interference by both CK-MM and CK-BB. Labeling anti-CK-MB with acridinium ester and immobilizing anti-CK-BB on paramagnetic particles, we developed a rapid and highly sensitive chemiluminescent/magnetic separation CK-MB assay. As little as 1 microgram of CK-MB per liter was detectable after 10- or 30-min incubation at room temperature, and the standard curve was linear up to 400 micrograms/L. Results for serum samples by the new assay correlated well (r = 0.94) with those by Corning electrophoretic and the Hybritech Tandem-E immunoenzymometric CK-MB methods. Sera containing macro CK-1 or high concentrations of CK-MM and CK-BB did not interfere. The combined advantages of a more-specific antibody, paramagnetic solid phase, and chemiluminescent label endow this two-site CK-MB assay with performance characteristics and ease of use superior to those of previous assays.     

At what level of serum total creatine kinase activity can measurement of serum creatine kinase MB isoenzyme activity be omitted in suspected myocardial infarction?

The purpose of this study was to establish a discriminatory limit for serum total creatine kinase activity (CK activity) below which CK isoenzyme fractionation is unnecessary. We looked at 2610 serum samples from 1077 consecutive patients with suspected acute myocardial infraction (AMI). The CK activity was determined according to the Scandinavian recommended method. Isoenzymes of CK were separated by agarose gel electrophoresis, followed by fluorometric scanning. When the threshold for CK activity was 150 U/l, none of the samples had a creatine kinase MB isoenzyme activity (CK-MB activity) equal to or higher than 30 U/l (the diagnostic level), which has been found to differentiate between patients with AMI and those without AMI. Only 14 patients (1.3% of all patients investigated) had CK-MB activity peaks between 10 U/l (detection limit) and 30 U/l. Of these, AMI was only diagnosed in one. We recommend that CK-MB activity should be measured only when CK activity is higher than 150 U/l. This would make about 50% of all CK-MB measurements unnecessary.     

Mass concentration and activity concentration of creatine kinase isoenzyme MB compared in serum after acute myocardial infarction

We compared three current methods (immunoinhibition, "Isomune-CK" immunoprecipitation, and the Tandem-E CKMB II immunoenzymometric assay) for determination of creatine kinase (CK; EC 2.7.3.2) isoenzyme MB in serum. Although results inter-correlated well, the immunoinhibition assay gave higher activity values. Atypical CK forms did not interfere with the immunoprecipitation and immunoenzymometric methods. In acute myocardial infarction the catalytic properties of CK decreased with the enzyme's age, as reflected by a steady increase in activation energy of the catalyzed reaction. In septicemia patients with very low CK and CK-MB catalytic activity, mean CK-MB mass concentration exceeded the upper reference limit, suggesting an increased rate of loss of activity concentration in these patients' sera. Because of the assay's lesser susceptibility to conformational changes at the active site of the enzyme, we suggest that measurement of CK-MB mass concentration is better suited for infarct sizing than measurement of catalytic activity.