Serum enzymes (aspartate transaminase, alanine transaminase, alkaline phosphatase (ALP), gamma-glutamyltransferase, and creatine kinase (CK] were measured in 296 young persons who admitted to recent inhalation of solvents, usually toluene based glues. In general, results fell within expected adult reference ranges except for ALP and CK. About 60% of subjects had CK activities above the upper reference limit and these activities were investigated in terms of their isoenzyme composition. CK B subunit activity was measured in 90 subjects with raised total CK activities. In five instances the CK B subunit activity was judged abnormal and in two subjects the presence of CK BB was confirmed. These two subjects were thought to have a circulating macro CK, type 1. It is concluded that the increased total CK activity found in this group of solvent abusers was due to physical activity, but a contribution from specific muscle toxicity by solvents cannot be excluded.

Patterns of creatine kinase (<em>CK</em>, EC 2.7.3.2) isoenzymes were studied in apparently healthy one- to 10-day-old neonates, by use of a sensitive fluorescent staining method with Sclavo <em>CK</em>-F/6001 reagent. Mean activities of <em>CK</em>3 (MM, 105 U/L), <em>CK</em>2 (MB, 6.8 U/L), <em>CK</em>1 (<em>BB</em>, 11 U/L), adenylate kinase (EC 2.7.4.3) anodal to <em>CK</em>3, and a fluorescent albumin artifact were found. Pooled plasma from neonates is recommended as a control because it defines the albumin artifact and approximates the activity of <em>CK</em>2 that must be observed after proper staining before a diagnosis of myocardial infarction can be made.

The MM, MB and BB isoenzymes of human creatine kinase (CK) were separated by elution from micro-columns of DEAE-Sephadex A-50 with Tris buffer containing increasing concentrations of NaCl at pH 7.0, instead of pH 8.0 as has commonly been used. Since pH 7.0 is close to the pH optimum of CK, this allowed the use of four times larger aliquots of the eluates for the estimation of CK activity and, consequently, a 4-fold increase in sensitivity. Using serum specimens from patients with acute myocardial infarction, there was a good correlation of the CK-MM (r = 0.99) and CK-MB (r = 0.93) activities obtained with the two buffer systems. Similarly, normal sera had CK-MB and CK-BB activities of less than 2 U/l with both buffer systems. Comparison of the composition of serum proteins in the eluates by conventional electrophoresis revealed that although the distribution of CK isoenzymes separated by the two buffer systems was similar, the distribution of proteins at pH 7.0 showed an appreciable shift of protein from the MB to the MM eluates.

WE HAVE DEVELOPED THREE TYPES OF EXPERIMENTAL SYSTEMS FOR THE STUDY OF SCCL: (1) serially heterotransplanted tumors in athymic nude mice; (2) continuous, clonable cell cultures; and (3) direct clonogenic assays for tumor specimens. These systems have their own individual advantages, applications, and limitations, but these are interrelated and complementary. The study of these systems has greatly aided our understanding of the biology of SCCL, and its relationship to other lung cancers and the APUD cell system. In addition, new markers for SCCL have been identified, such as a creatine kinase and its BB isoenzyme (CK-BB). These cellular markers may have clinical applications, as serum levels of CK-BB are an indicator of tumor burden. Assays for clonogenic tumor cells may permit selection of optimal drug combinations for the treatment of individual tumors. Variant cultures having the morphology of SCCL, but lacking some or all of the other features, have been identified. While our systems have been used primarily for biological studies, they have clinical applications for both diagnostic and therapeutic purposes.

A new commercial kit (Impres-MB; International Immunoassay Labs.) recently was introduced for measuring the MB isoenzyme of creatine kinase (CK-MB) based on the use of monoclonal antibodies. After antibodies to CK-MM isoenzyme are added to precipitate the CK-MM, antibodies to CK-M monomer are added to precipitate the M-subunit isoenzymes of CK. Subtracting the enzymatic activity of the second supernate from the residual activity in the first yields the activity of CK-MB. Results are not affected by CK-BB, mitochondrial CK, or adenylate kinase. However, the anti-CK-MM antibodies precipitated only about 98% of serum CK-MM and may have partly precipitated CK-MB isoenzyme (average analytical recovery of CK-MB, 86.6%). Comparison between Impres-MB (y) and electrophoresis (x) yielded the following linear-regression equation: y = 0.79x + 3 (r = 0.982, n = 97). Data for CK-MB temporal kinetics, obtained from patients with myocardial infarction, correlated significantly in both methods; however, peak activity values of CK-MB were significantly different, confirming that the difference between the new method and the electrophoretic method average 20%.

A total of 979 cardiac profiles were reviewed. Seventeen cases were found to have elevated CK-BB by electrophoresis and were misidentified by the immunoinhibition/immunoprecipitation technique as elevated creatine phosphokinase (CK-MB). Eleven of the 17 cases also had elevated lactate dehydrogenase (LD) LD-5/LD-1 ratio; five cases were motor vehicle accident (MVA), four cases were prostatic carcinoma (PC), and one case each of breast carcinoma and coronary heart disease. One case of PC and one of MVA with a preliminary clinical diagnosis of acute myocardial infarction (AMI) were presented. Our findings underscore the importance of electrophoretic confirmation of the presence of CK-MB when detected by a quantitative technique. Clinicians should consider the possibilities of PC or other cancers when elevated "CK-MB" is present in conjunction with a raised LD-5/LD-1 ratio in patients who fail to show clear-cut clinical evidence of AMI. The mechanism of elevated CK-BB and LD-5/LD-1 ratio in PC patients are discussed.

In this "column-batch" method for separating the MB and BB isoenzymes of creatine kinase and the LD1 isoenzyme of lactate dehydrogenase, one can, alternatively, separate MB from BB or obtain a combined fraction containing MB, BB, and LD1. The principal advantage is that the resulting fractions are twofold as concentrated as was the applied sample. Thus, activity can be measured by conventional automated methods, with no need for the modifications to compensate for diluted fractions that are required by other ion-exchange methods. Another advantage is the total absence of interference by the MM isoenzyme. A strong anion exchanger (AG-MP1, Bio-Rad) is used in the acetate form at pH 6.3. There is no retention of MM; retained MB, BB, and LD1 are eluted with a solution of magnesium acetate. Results are compared with those obtained for subunit B and LD1 by immunoinhibition. Results with patients are considered consistent with myocardial infarction if MB exceeds 20 U/L and 3% of the total CK and LD1 exceeds 130 U/L or 28% of the total LD activity.

The purpose of this study is to determine the levels of brain type isoenzyme of creatine kinase (CK-BB) as a possible indicator of a pre-existing intrauterine brain-cell damage in cord blood sera of fetuses with preceding absent or reverse end-diastolic flow velocities of the umbilical arteries (AREDFV). CK-BB isoenzyme activities were determined in umbilical cord sera of 13 newborn infants with preceding AREDFV and in 14 fetuses with low end-diastolic flow velocities (LEDFV) of the umbilical arteries. 50 newborn infants with elective cesarean section and normal umbilical artery blood flow velocity waveforms were used as controls. Two-tailed Student's t-test and Fischer's exact test were used for statistical evaluation of the results. CK-BB isoenzyme activity did not depend on gestational age. Fetuses with AREDFV showed a significant increase in CK-BB values, whereas fetuses with LEDFV had CK-BB activities within the normal range of the controls. The elevated CK-BB values of the AREDFV group were not correlated with fetal acidosis at birth. Brain-cell injury with leakage of CK-BB isoenzyme might be present in fetuses with AREDFV even before (preterm) delivery.

Hepatocyte transplantation has been explored as a therapeutic alternative to liver transplantation, but a means to monitor the success of the procedure is lacking. Published findings support the use of in vivo (31)P MRSI of creatine kinase (CK)-expressing hepatocytes to monitor proliferation of implanted hepatocytes. Phosphocreatine tissue level depends upon creatine (Cr) input to the CK enzyme reaction, but Cr measurement by (1)H MRS suffers from low signal-to-noise ratio (SNR). We examine the possibility of using the Cr analog cyclocreatine (CCr, a substrate for CK), which is quickly phosphorylated to phosphocyclocreatine (PCCr), as a higher SNR alternative to Cr. (1)H MRS and (31)P MRSI were employed to measure the effect of incremental supplementation of CCr upon PCCr, γ-ATP, pH and Pi /ATP in the liver of transgenic mice expressing the BB isoform of CK (CKBB) in hepatocytes. Water supplementation with 0.1% CCr led to a peak total PCCr level of 17.15 ± 1.07 mmol/kg wet weight by 6 weeks, while adding 1.0% CCr led to a stable PCCr liver level of 18.12 ± 3.91 mmol/kg by the fourth day of feeding. PCCr was positively correlated with CCr, and ATP concentration and pH declined with increasing PCCr. Feeding with 1% CCr in water induced an apparent saturated level of PCCr, suggesting that CCr quantization may not be necessary for quantifying expression of CK in mice. These findings support the possibility of using (31)P MRS to noninvasively monitor hepatocyte transplant success with CK-expressing hepatocytes.

The concentration of brain type creatine kinase (CK-BB) was measured in blood from the internal jugular vein in 32 children (less than 1 year old) with congenital heart disease. In transposition of the great arteries the CK-BB levels were significantly higher than in children without cyanosis (10.1 +/- 4.1 vs. 3.0 +/- 0.5 ng/ml). A negative correlation was found for CK-BB concentration and arterial oxygen saturation (r = -0.41, p less than 0.02 for all children and r = -0.62, p less than 0.05 for those with tetralogy of Fallot). It is suggested that the increased CK-BB levels in the blood of cyanotic children reflect chronic cerebral hypoxia, which may explain other reports of reduced psycho-intellectual function in patients with cyanotic heart disease.

During the last three years we and other have observed discrepancies between results for creatine kinase isoenzyme MB as measured with the mechanized ion-exchange chromatographic method in the Du Pont aca and those by other techniques. These observations prompted us to investigate the influence of the matrix on the Du Pont CK-MB assay. We conclude that, apart from possible interferences by CK-MM, CK-BB, and both types of macro CK, the aca will give apparent CK-MB activities that are directly related to protein concentration and inversely related to sodium chloride concentration. Practical consequences for the routine and emergency laboratories are: no diluted samples are allowed; application is restricted to samples from patients suspected of acute myocardial infarction which show upper-normal total CK activity; and multiple timed samples are run, in order to recognize the typical change in enzyme pattern with time.

The association between measurements of lateral ventricle dilatation determined by serial ultrasound and brain specific creatine-kinase isoenzyme patterns (CK-BB) is studied in 60 very low birth weight preterm neonates of 1,500 g birth weight or 32 weeks gestation or less. The patients were divided into three groups according to cranial ultrasonographic findings: Group A (n = 20) had isolated peri-intraventricular hemorrhage (PIVH); group B (n = 20) had PIVH and dilated ventricles (VM); group C (n = 20) were normal matched preterms and formed the control group. Compared to control babies or those with isolated PIVH, high serum concentrations of CK-BB were observed after birth in babies with persistent dilated ventricles at two weeks postnatal age (p less than 0.01). No difference was found between CK-BB levels of babies with isolated PIVH and control group (p greater than 0.05). We suggest that an elevated CK-BB value is found in babies with persistent ventricular dilatation suggesting severe and diffuse brain damage after post-hemorrhagic ventriculomegaly (VM).

A study was made of the content of creatine kinase-BB (CK-BB) and lactate in cerebrospinal fluid (CSF) of 202 neonates and infants with perinatal CNS injuries. The relationship was found between the rise of the CK-BB content and the gravity of perinatal CNS injuries. The highest content of CK-BB in CSF was marked in neonates with cerebral disorders complicated by infectious and inflammatory diseases (pneumonia, sepsis). Within the first 5 days of life, the children of this group demonstrated the relationship between the content of CK-BB and lactate of CSF. The measurement of the content of CK-BB in CSF should be used for early diagnosis, assessment of the gravity and course of perinatal CNS injuries in neonates and in infants.

Profound hypothermia and circulatory arrest in infants and children undergoing cardiac surgery were followed by abnormally high plasma levels of creatine kinase isoenzyme BB (CK-BB). Differences in the levels of enzymes in the femoral arterial and jugular venous blood indicated that the origin of the additional enzyme was the brain. The extent of the rise in enzyme levels was related to the duration of circulatory arrest. These data suggest that measurements of the CK-BB enzyme in plasma provide quantitative information about cerebral damage during cardiac surgery.

The clinical value of serum brain specific creatine kinase (CK-BB) was assessed in head injured patients (group A) using a new enzyme-linked immunosorbent assay (ELISA). The results were compared to healthy controls (group B) and patients post-myocardial infarction (group C). None of the head injured patients had undergone a surgical procedure or ventricular puncture. CK-BB was significantly higher in group A than in controls. The level of CK-BB in group A was inversely proportional to the Glasgow Coma Scale on admission. All patients with a CK-BB greater than 100 micrograms/l died. The ELISA technique is a simple and reliable assay with prognostic significance in patients with head injury and has wider clinical application than the previously described radioimmunoassay methods.

Creatine kinase (CK) isoenzymes are considered to be specific tissue injury markers. The purpose of this study was to evaluate CK isoenzymes (CK-MM, CK-MB, and CK-BB) in umbilical cord blood sera of newborns in relation to their acid-base status. Investigations were performed in a group of 75 newborns delivered after 37 weeks of pregnancy. We estimated CK-isoenzymes with the use of DEAE-Sepharose C1-6B column chromatography. Total CK activity was measured using kits supplied by Boehringer-Mannheim (Monotest-R CK-NAC aktiviert). Newborns were considered hypoxic if umbilical artery blood pH was under 7.2 and/or base excess less than -10 mmol/L. Of 75 newly born infants 26 had features of perinatal hypoxia. We were unable to demonstrate significant differences in total CK, CK-MM and CK-MB activities in examined groups of newborns. We found a significant rise of CK-BB activity in cord sera of hypoxic infants. Our results show that the fetal brain is the most vulnerable to a hypoxic state.

A diagnosis of myocardial infarction (MI) is usually established by the evaluation of clinical symptoms, electrocardiographic changes, and serum enzyme levels, specifically creatine phosphokinase, subunit MB (CK-MB), by electrophoresis. A total of 215 patients were evaluated in this study. One hundred two of them were admitted to the coronary care unit and 113 to the emergency room, where they were screened for possible MIs. The radioimmunoassay (RIA) used in this study determines levels of the CK-MB isoenzyme by detecting the B monomer, which also has 100% cross-reactivity with the CK-BB isoenzyme. The intra-assay coefficients of variability (CVs) for 30 samples were 22% (means = 7.0 ng/ml) and 11% (means = 47.3 ng/ml), and the interassay CVs for 30 samples were 17% (means = 7.1 ng/ml) and 9.2% (means = 49.3 ng/ml). Of the 215 patients evaluated, 21 had myocardial infarction by the criteria in the study. The diagnostic sensitivity, specificity, and accuracy were 100.0%, 92.8%, and 93.5% respectively. These values increased to 100.0%, 96.9%, and 97.2% when only coronary care unit patients were considered. The CK-MB RIA was found to be a reliable replacement for electrophoresis, but it was nonspecific in some patients.

The value of radioimmunoassay for creatine kinase BB isoenzyme (CK-BB) determination in early diagnosis of acute myocardial infarction was estimated. The clinical material consisted of 35 randomly selected patients admitted to a coronary care unit during the 4 h after chest pain suspected for myocardial infarction. In all patients standard 12 lead electrocardiograms were obtained on admission and at 24, 48 and 72 hours after admission. Blood samples for CK-BB analysis were collected on admission and at 4 hourly intervals for 48 h after admission. Aspartate aminotransferase (GOT) activity was determined in all patients on admission and in samples obtained at 24 and 48 hours. The patients were classified to 3 groups according to electrocardiographic and clinical findings. The first group consisted of 10 patients whose electrocardiograms fulfilled the criteria of transmural infarction. The electrocardiograms of 10 patients of the second group fulfilled the criteria of subendocardial infarction. The remaining 15 patients made up a third group of coronary insufficiency. The activity of CK-BB at various time intervals after chest pain in all groups was compared in order to estimate the value of this method for differentiation of the 3 causes of coronary pain. The frequency of positive results (the values exceeding upper limits of normal range for healthy people) at various time intervals after pain obtained with both enzymatic methods was compared in order to estimate the sensitivity of analysed method.(ABSTRACT TRUNCATED AT 250 WORDS)

We describe the case of an elderly woman whose symptoms and electrocardiographic pattern initially suggested acute myocardial infarction. The value for total serum creatine kinase (EC 2.7.3.2; CK) was 737 U/L (reference interval: 22-269 U/L), and electrophoresis for CK isoenzymes demonstrated two bands, the more anodal migrating to the CK-MB region and the second migrating between the CK-MB and CK-MM regions. The above-normal total CK and electrophoretic pattern persisted during her 11-day hospital course. The QuiCK-MB (International Immunoassay Labs.) and Tandem-E CK-MB (Hybritech) immunoassays, however, showed CK-MB mass measurements within the normal range. In further investigation with a mixture of patient's serum and human-serum-based control containing all CK isoenzymes, the electrophoretic mobility of only CK-BB was altered, proving that the patient had antibody to the B unit of CK in her serum. Immunofixation revealed the more anodal band to be a CK-IgA lambda complex, and the more cathodal band, a CK-IgG kappa complex. Mixing the patient's serum with polyclonal antibody specific for CK-B slowed the electrophoretic mobility of only the more anodal band. Polyclonal antibody specific for CK-M had no effect on either band. Evidently, this patient had two different types of macro CK type 1, both containing CK-BB. We conclude that macro CK type 1 can mimic CK-MB and be a source of confusion.

The clinical usefulness of the Tandem-E CK-MB method (A) using two monoclonal antibodies to measure the intact cardiac specific creatine kinase isoenzyme (CK-MB) was evaluated against results obtained by Corning's electrophoretic fluorescence scanning method (B) and the DuPont aca mechanized column chromatographic method (C). In 95 patients suspected to have myocardial infarction, a total of 127 CK's with MB's were simultaneously determined by methods "A", "B", and "C". "C" produced a total of 16.5% (21 out of 127) false-negative or false-positive results. In contrast, CK-MB results obtained by "A" correlated very well with those determined by "B" (r = 0.97; N = 64). In addition, the clinical course of myocardial infarction as monitored by the measurement of CK-MB at various time intervals by both methods showed excellent parallelism. Furthermore, "A" was shown to be free of interference from the presence of macro CK type I and II. Also, the presence of increasing concentrations of CK-BB (up to 29 IU/L) did not alter the assay response of CK-MB in these serum samples. In conclusion, we found the Tandem-E CK-MB method to be sensitive (sensitivity = 94%) and specific (specificity = 96%).

The activity of creatine kinase isoenzyme BB (CK-BB) in serum is rarely abnormally high (i.e., detectable). An increase in immunoreactive CK-BB or CK-BB activity in patients with prostatic disease has been proposed as an indication of prostatic adenocarcinoma. Here we report the case of an elderly man with massive benign prostatic hyperplasia but no clinical or pathological evidence of prostatic adenocarcinoma, whose serum CK-BB activity was found by agarose gel electrophoresis to be 1 U/L (normal: 0%), 10% of his total CK activity. Serum CK-BB activity was further increased to 16 U/L (20% of total CK activity) 1 h after prostatectomy, but became undetectable by the second day after the operation. The findings suggest that: (a) the source of the serum CK-BB activity was the enlarged prostate gland; (b) abnormally high CK-BB activity in serum of men with prostatic disease does not necessarily indicate the presence of prostatic adenocarcinoma; and (c) myocardial injury could be erroneously diagnosed postoperatively in prostatectomy patients if CK isoenzyme methods are used that do not consistently separate "heart-specific" CK-MB from CK-BB.

We examined the clinical and analytical performance of two immunoassays (Becton Dickinson CK-MB; Ciba-Corning Magic Lite CK-MB) in which monoclonal anti-CK-MB antibodies are used for directly measuring creatine kinase (EC 2.7.3.2) isoenzyme MB (CK-MB) in serum, and also one electrophoretic method (Ciba-Corning). Within- and between-assay precision for both immunoassays was good at the upper reference limits (less than 10% CV). Analytical recoveries ranged from 102 to 114%. Both immunoassays were free from interference by CK-BB, mitochondrial-CK, macro-CK, adenylate kinase, and CK-MM. Retrospectively, we evaluated four categories of patients, using both immunoassays and electrophoresis: normal controls, acute myocardial infarction (AMI) patients, severe skeletal muscle trauma patients, and acutely ill patients known not to have AMI. In general, there were excellent correlations among all three methods. CK-MB activity (U/L) measured by the Becton Dickinson immunoassay was approximately 50% of the mass concentration (microgram/L) of the Magic Lite immunoassay and 50% of the activity concentration (U/L) determined by electrophoresis. Both immunoassays were easy to perform and sensitive to the low CK-MB concentrations often found with low total-CK activities.

Recent investigations have shown that cardiac isoenzymes change with mechanical overload and possibly with myocardial ischaemia. This complicates the interpretation of serum enzyme changes in acute myocardial infarction. We have therefore investigated the rate of release of isoenzymes from necrosing myocardium and the effect of ischaemia per se. Discrete myocardial infarction was produced in 35 male Wistar rats by ligation of left coronary artery. Six (n = 7), 12 (n = 6), 24 (n = 9), 72 (n = 7) h and 3 weeks (n = 6) after surgery, total and isoenzyme activities of creatine kinase (CK), lactate dehydrogenase (LD) and aspartate aminotransferase (AST) were measured in the infarcted myocardium. Untreated rats (n = 12) were used as the control (time 0). Sham operation was performed in 36 rats. During the early period (0 to 12 or 24 h) of infarction, each (iso)enzyme disappeared monoexponentially from the myocardium (mean r = 0.88) with different disappearance rates. Cytosolic isoenzyme fractions decreased more rapidly than mitochondrial fractions. CK MB and the LD-H subunit decreased faster than CK MM and the LD-M subunit. Such differences in the disappearance rate may be related to subcellular localisation of each isoenzyme. In the late period (72 h and 3 weeks), CK BB and the LD-M subunit showed significant reaccumulation in the infarcted myocardium. Although inflammatory cells can be responsible for the reaccumulation of LD-M subunit, the origin of CK BB is unknown.(ABSTRACT TRUNCATED AT 250 WORDS)

OBJECTIVE

To investigate the relationship of creatine phosphokinase and its isoenzymes with fetal asphyxia and risk at birth.

METHODS

Thirty-five pregnant women with high-risk pregnancy were studied.

RESULTS

In 21 patients, fetal distress was diagnosed by interpretation of the fetal heart rate tracing (FHR). The remaining 14 women, having normal fetal cardiotocography, were considered as the control group. Total CK and its isoenzymes activity was measured in cord sera and 24 h after birth in peripheral blood. Abnormal FHR patterns correlate well with elevated enzyme activities. Total CK and its isoenzymes (CK-MM, CK-MB, and CK-BB) exhibited higher values in asphyxiated infants as compared to normal neonates. Electrocardiographic ischemia occurred in seven newborns who had elevated CK-MB and CK-BB levels, both at birth and within 24 h postpartum. Chromatographic study showed in normal neonates that the predominant isoenzyme was CK-MM, whereas CK-BB activity was negligible. In the newborns with abnormal FHR, CK-MB and CK-BB were increased with predominance of CK-MB.

CONCLUSIONS

Antepartum fetal distress is associated with release of CK-BB, and particularly CK-MB; therefore, these biochemical markers may indicate either brain or myocardial damage.