Immunoenzymetric assay for creatine kinase MB with subunit-specific monoclonal antibodies compared with an immunochemical method and electrophoresis

Results of an immunoenzymetric assay (TANDEM-E CKMB) for creatine kinase (CK; EC 2.7.3.2) MB isoenzyme, in which subunit-specific monoclonal antibodies are used, were compared with those by an immunochemical method (Isomune-CK) and electrophoresis (Corning agarose gel). The study involved 200 patients; greater than 500 samples were analyzed by all three methods. The analytical performances were acceptable. Between-method correlation coefficients ranged from 0.881 to 0.975. Two reference intervals were established for the immunoassays: 0-4 micrograms/L (TANDEM) and 0-4 U/L (Isomune) for "normal" patients; 0-9 micrograms/L (TANDEM) and 0-14 U/L (Isomune) for noninfarct patients. Agreement with respect to increased CK-MB as defined by the reference intervals for the noninfarct patient was 96% between TANDEM and electrophoresis, 90% between Isomune and electrophoresis. All three methods are acceptable for use in determining CK-MB, but the overall diagnostic efficiencies for the mass or activity concentration of the isoenzyme and for its proportion of total CK activity, based on the predictive value model, are 92% (electrophoresis, 0-7 U/L), 90% (electrophoresis, 0-4%), 92% (TANDEM, 0-9 micrograms/L), 88% (TANDEM, 0-3% index), 88% (Isomune, 0-14 U/L), and 83% (Isomune, 0-4%). All three methods can detect CK-MB in serum, but its presence is not necessarily diagnostic of acute infarct. We recommend using the actual concentration of CK-MB to evaluate patients with suspected acute myocardial infarct, and the percentage of CK-MB when total CK is very high.     

Clinical and analytical evaluation of two immunoassays for direct measurement of creatine kinase MB with monoclonal anti-CK-MB antibodies

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.     

Diagnostic efficacy of a new enzyme immunoassay for creatine kinase MB isoenzyme

A new commercial enzyme immunoassay kit for quantification of creatine kinase-MB (CK-MB) isoenzyme was compared with its electrophoretic determination with respect to efficacy in diagnosis of acute myocardial infarction. Enzygnost CK-MB (Behring Diagnostics) is a solid-phase "sandwich"-type enzyme immunoassay with antibodies to the B-subunit coated on plastic tubes and peroxidase-conjugated antibodies to the M-subunit added after incubation with sample. This kit is designed to measure only CK-MB and not CK-MM, CK-BB, adenylate kinase, or atypical CK molecules. The linear-regression equation comparing the two methods was: Enzygnost = 0.98 . electrophoresis - 0.72, with a correlation coefficient of r = 0.967 (n = 143). For 51 patients admitted for diagnosis of possible acute myocardial infarction, the Enzygnost kit achieved 100% sensitivity, specificity, and efficiency in predicting the correct diagnosis. Corresponding values for the electrophoretic assay were: 95.5% sensitivity, 93.1% specificity, and 94.1% efficiency. We conclude that this kit method provides an excellent alternative to electrophoresis.     

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.     

Evaluation of a commercial immunoenzymometric assay kit for creatine kinase MB isoenzyme determination using monoclonal antibodies

A simultaneous two-site immunoenzymometric assay for creatine kinase MB determination (Hybritech Tandem-E CK-MB) using monoclonal antibodies was evaluated and compared with cellulose acetate electrophoresis using fluorometric scanning densitometry. The assay has satisfactory precision (between-day analysis gives a coefficient of variation between 2.1 and 9.4%) and is not susceptible to interference by concentrations of creatine kinase MM up to 5000 micrograms/l (3400 U/l) and creatine kinase BB up to 1000 micrograms/l (1085 U/l). The upper limit of MB isoenzyme concentration in 250 apparently healthy people was 5.5 micrograms/l. Comparison between the immunoenzymometric assay (y) and electrophoresis (x) yielded the following linear regression equation: y = 0.37x + 1.9, with a correlation coefficient of 0.828. The characteristics of the temporal kinetics of MB isoenzyme, calculated by two methods, in 49 patients with acute myocardial infarction, were nearly identical in terms of the rate of creatine kinase MB release and the time at which the peak value is obtained, but not in terms of the rate of elimination of the isoenzyme. The fractional disappearance rate of MB isoenzyme from the circulation was significantly higher if calculated with Tandem-E results rather than with electrophoresis results (-0.035 vs -0.028, p less than 0.001). Whereas in the first day after infarction immunoenzymometric assay and electrophoresis had the same clinical sensitivity for identifying patients with acute myocardial infarction, in specimens collected more than 24 hours after the onset of the chest pain, the clinical sensitivity of the immunoenzymometric method was lower. Our results show that it is still premature to draw definitive clinical conclusions from the immunoassay results.     

Concordance of creatine kinase-MB activity and mass

The recent availability of monoclonal antibodies that are highly specific for creatine kinase (CK; EC 2.7.3.2) MB isoenzyme should allow for the development of rapid, sensitive, and specific assays of CK-MB mass and activity. However, the relationship between the mass concentration of CK-MB and its activity in plasma has previously been thought by some to be variable. To determine the extent to which discrepancies of potential clinical significance might arise between measurements of activity and mass in plasma, we compared CK-MB activity and concentration in 1298 samples obtained from 226 patients admitted to the cardiac-care unit. CK-MB activity concentration was determined with an immunoadsorption assay, and mass concentration was measured by an automated "sandwich" assay (Magic Lite; Ciba Corning Diagnostics). Both of these assays are based on specific monoclonal antibodies for CK-MB. Values obtained with these assays correlated well (r = 0.94). Normal and abnormal values with the two assays were concordant in 96% of the samples. In all but three instances, differences occurred late after myocardial infarction and were characterized by minimal increases as determined by one method vs values at the upper limit of normal as determined with the other. Thus, measurements of CK-MB mass and activity concentrations in plasma with assays based on these specific monoclonal antibodies are comparable for the detection or exclusion of acute myocardial infarction.     

Clinical and analytical evaluation of different methods for measurement of creatine kinase isoenzyme MB

We evaluated the clinical and analytical performance of the new immunochemiluminometric assay (ICMA; Ciba Corning) for measurement of creatine kinase isoenzyme MB (CK-MB), and compared it with three other methods: immunoradiometric assay (IRMA; International Immunoassay Labs); immunoinhibition assay (Seradyn); and an immunoinhibition/column method (Du Pont). Intra-test precision for all kits was good. We evaluated 32 patients' samples by all four methodologies. Only one of the four methods (aca, Du Pont) showed evidence of linearity. Efficiency in the diagnosis of myocardial injury in our study ranged from 53% (Seradyn) to 96% (Du Pont). We evaluated serial specimens from 20 separate patients by the IRMA and the ICMA to determine whether myocardial injury could be diagnosed earlier by the ICMA. In patients with acute myocardial infarction, the ICMA displayed positive values earlier and longer than the IRMA, suggesting that the ICMA is suited for screening for myocardial damage in hospitalized patients.     

Analytical and clinical evaluation of creatine kinase MB mass assay by IMx: comparison with MB isoenzyme activity and serum myoglobin for early diagnosis of myocardial infarction.

We analytically and clinically evaluated Abbott's IMx assay for creatine kinase (CK) isoenzyme MB (CK-MB) in serum. Over a 1-year period, the method was more specific but less precise than catalytic isoenzyme measurements by electrophoresis or immunoinhibition. Sera from different individuals without electrophoretic evidence of CK-MB but containing macro CK type 1 (n = 20), mitochondrial CK (n = 5), or CK-BB (n = 5) were scored as CK-MB negative by the IMx. Likewise, CK-MB-negative by the sera remained so after addition of purified human CK-MM (< or = 7600 U/L) or CK-BB (< or = 8100 U/L). For 39 patients admitted for suspicion of uncomplicated acute myocardial infarction (precordial pain for < or = 4 h), the diagnostic performance of the IMx CK-MB assay on admission and 4 h later was superior to that of total CK activity and compared well with that of CK-MB activity measured by electrophoresis or immunoinhibition. An admission, myoglobin showed a higher diagnostic sensitivity, specificity, and predictive value than did CK-MB and was the most informative test. Diagnostic performance on admission and 4 h later was further improved by considering positivity for myoglobin and for CK-MB by IMx and for the change in each over the first 4 h of hospitalization as criteria. Twelve hours after admission, diagnostic performance was further improved for all CK and CK-MB methods but began to decline for myoglobin.     

Semi-automated direct colorimetric measurement of creatine kinase isoenzyme MB activity after extraction from serum by use of a CK-MB-specific monoclonal antibody

This semi-automated colorimetric assay for the MB isoenzyme of creatine kinase (EC 2.7.3.2) is based on a monoclonal antibody ("Conan-MB") specific for this isoenzyme and is a modification of a previously published method (Vaidya et al., Clin Chem 1986;32:657-63). A 0.64-cm bead coated with 2 to 3 micrograms of antibody is incubated with 100 microL of serum and 10 microL of 0.2 mol/L beta-mercaptoethanol for 1 h at room temperature, to extract CK-MB. The beads are washed with de-ionized water and incubated with CK substrate for 45 min at 37 degrees C. A solution containing trans-1,2-diaminocyclohexane-N,N,N', N'-tetraacetic acid, p-iodonitrotetrazolium violet, and diaphorase is added and the resulting colored product is measured at 492 nm. The standard curve is linear to 200 U of CK-MB per liter, and analytical recovery is 97-113%. Total assay CV for low (9.7 U/L) and high (50.7 U/L) quality-control materials was 14.1% (n = 1878) and 11.6% (n = 1842), respectively. CK-MB activity correlated well (r = 0.978, n = 226) with CK-MB measured by a two-site mass immunoassay, and 99.4% of samples with CK-MB greater than or equal to 12 U/L (n = 347) were verified by electrophoresis on agarose.     

Influence of autoantibodies to creatine kinase-BB on assays for MB isoenzyme

We describe the influence of autoantibodies that bind creatine kinase BB (CK-BB) on the methods for MB isoenzyme. If these autoantibodies are present in patients' sera, they cause the formation of macro CK type 1 (immunoglobulin-linked CK-BB). In some of these cases they can bind not only endogenous CK-BB but also CK-MB without significantly affecting enzyme activity. Although these antibodies show distinctly less affinity for CK-MB than for CK-BB, they nevertheless bind CK-MB in these particular sera, because their concentration exceeds that of CK-BB isoenzyme. If a person with such autoantibodies has an acute myocardial infarction, the immunoinhibition method for CK-MB, which does not discriminate between CK-MB and CK-BB, will recognize the increase and peak of CK-MB with time, although persistent macro CK activity will be superimposed on the typical isoenzyme pattern. However, isoenzyme electrophoresis and recently introduced immunoenzymometric assays for CK-MB in these cases may be less sensitive for detecting myocardial infarctions, because the typical increase in CK-MB activity may be identified later in the progression of symptoms, or even be missed.     

Creatine kinase isoenzyme MB assay by electrophoresis

A method for determination of the creatine kinase isoenzyme MB (CK-MB) is reported: separation of the isoenzymes was done by electrophoresis and the activity of the isoenzyme bands quantitated by scanning fluorometry. Total CK activity was used for calculation of CK-MB level. The precision of the method was satisfactory: coefficient of variation 5-10%. Its accuracy good: CK-MB was consistently found in high concentrations in tissue extracts of myocardium, but was virtually absent in skeletal muscle and could not be demonstrated in serum from patients with skeletal muscle damage. The sensitivity of the method fitted its clinical use: CK-MB was undetectable (less than 5 U/l) in normal sera, below 30 U/l in seventy-six out of seventy-seven patients in whom the diagnosis of acute myocardial infarction (AMI) was disproved, and above 30 U/l in all seventy-two patients with AMI according to WHO criteria. The CK-MB concentration in serum rises to a maximum about 20 h after onset of clinical symptoms of AMI and reaches baseline levels 20-30 h later. The electrophoretic CK-MB method is easy, fast and reliable and is considered as an important diagnostic test for AMI.     

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.     

Evaluation of the clinical usefulness of a chemiluminometric method for measuring creatine kinase MB

The use of creatine kinase isoenzymes (CK-MB) in the diagnosis of acute myocardial infarction (AMI) is well established. We evaluated the use of a new chemiluminometric method (CK-Ciba) for measuring CK-MB by calculating its sensitivity, specificity, positive and negative predictive values, and diagnostic efficiency for diagnosing AMI. We tested 633 samples from 229 patients within 4 h of receipt. The patients were divided into four groups: (1) patients who had an AMI, (2) patients who had AMI ruled out, (3) patients who had CK-MB measured for reasons other than to rule out AMI, and (4) patients who had only one sample drawn. Only patients in Groups 1 and 2 were used in the study. AMI was diagnosed by a cardiologist. The prevalence of AMI in our population was 0.18. A receiver-operator characteristic curve was used to establish optimal values for identifying AMI with the CK-Ciba results: CK-MB greater than or equal to 10 micrograms/L and a CK-MB index of greater than or equal to 3.0 (micrograms of CK-MB per U of CK x 100). Using these values, we calculated a sensitivity of 1.00, specificity of 0.97, positive predictive value of 0.87, negative predictive value of 1.00, and a diagnostic efficiency of 0.97. We conclude that the CK-Ciba method has high sensitivity, high specificity, and good predictive values for CK-MB and is appropriate to use to rule out AMI.     

Standardization of creatine kinase-MB (CK-MB) mass assays: the use of recombinant CK-MB as a reference material.

BACKGROUND: The AACC assembled a committee to identify and validate a standard creatine kinase MB isoenzyme (CK-MB) material to improve the comparability of CK-MB mass assays. METHODS: Three protocols were used. In protocol I, various CK-MB materials prepared in different matrices were screened as candidate standards. In protocol II, participating manufacturers calibrated their systems with concentrates of human heart CK-MB and then tested 20 patient samples to evaluate calibration bias. In protocol III, participating manufacturers calibrated their immunoassay systems using recombinant CK-MB2 (rCK-MB2) diluted into their respective sample diluents and measured 50 samples. RESULTS: Candidate materials showed high recovery in stripped human serum, but bias improved only from 59% to 38%. These data led to the use of human heart CK-MB diluted in each manufacturer's sample diluent. This strategy reduced bias from 31% to 15%. Because human heart CK-MB is difficult to provide, a lyophilized source of CK-MB2 was identified. rCK-MB2 was shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, reversed-phase HPLC, intrinsic protein fluorescence, circular dichroism, agarose gel electrophoresis, immunoreactivity studies, high and low temperature stability, and reconstituted stability to be equivalent to human heart CK-MB. Calibration of immunoassay systems with rCK-MB2 added into each respective manufacturer's sample diluent showed a 13% between-manufacturer bias. CONCLUSION: Lyophilized rCK-MB2 was determined suitable for use as a reference material for CK-MB mass assays.     

Activity concentration and mass concentration (monoclonal antibody immunoenzymometric method) compared for creatine kinase MB isoenzyme in serum

Results of the "Tandem-E CKMB" immunoenzymometric procedure (y) for creatine kinase (CK; EC 2.9.3.2) were compared with electrophoresis (x) for 160 serum samples from patients suspected of having sustained myocardial infarctions. The results correlated well: y, microgram/L (Tandem assay) = 1.3x-6.3 U/L(electrophoresis) (r = 0.95). CK-MB mass measurement was more stable than enzyme activity after storage and appeared to be more sensitive. Sera from 86 other people, which had no detectable CK-MB upon electrophoresis, gave a mean CK-MB value of 1.1 microgram/L (SD 1.3, range 0-8) with the Tandem assay. To determine whether these low values represented actual isoenzyme, we tested for possible interference by heterophile antibodies in the patients' sera by preincubating the samples with mouse serum before the Tandem assay. The mouse serum did not interfere with the assay of sera that had substantial quantities of CK-MB by electrophoresis. However, in five of six samples that were negative by electrophoresis, the CK-MB values were substantially smaller, indicating that the values measured were false-positives caused by the presence of heterophile antibodies directed against mouse proteins, an interference that could be eliminated by pretreatment with mouse serum.     

Comparison of enzyme immunoassay and immunoprecipitation for creatine kinase MB in diagnosis of acute myocardial infarction

We compared the clinical performance of measuring creatine kinase (EC 2.7.3.2) isoenzyme MB by use of an enzyme immunoassay (Enzygnost CK-MB, Behring Diagnostics) with an immunoprecipitation method (Isomune-CK, Roche Diagnostics) for the diagnosis of acute myocardial infarction. Sera from 80 patients admitted to the coronary care unit because of chest pain were examined: 40 who had this diagnosis of myocardial infarction, and 40 in whom it was ruled out. In addition, sera from 40 apparently healthy individuals were examined. The clinical sensitivity and specificity of these methods were evaluated by use of receiver operating characteristic curves. We conclude that for clinical efficiency, this enzyme immunoassay is slightly superior to the immunoprecipitation assay we used, because of its greater analytical sensitivity and precision for measuring the mass of the isoenzyme.     

Quantifying the MB isoenzyme of creatine kinase with the Abbott "IMx" immunoassay analyzer

In this two-step automated assay of the MB isoenzyme of creatine kinase (CK-MB), developed for the Abbott "IMx" immunoassay analyzer, monoclonal anti-CK-MB antibody immobilized onto latex microparticles and polyclonal anti-CK-MM antibody coupled to alkaline phosphatase are used. Within-run CVs ranged from 3.9% to 9.0%, between-run CVs from 0.0% to 5.6%, and the sensitivity was 0.2 microgram/L. Twenty-four results can be obtained in about 37 min. Analytical recovery of CK-MB added to human serum or plasma ranged from 89% to 109%. Icteric, lipemic, or hemolyzed samples did not interfere with CK-MB recovery. Cross-reactivity with CK-MM and CK-BB was 0.012% and 0.001%, respectively. The normal reference interval was 0-5 micrograms/L. IMx CK-MB results correlated well with CK-MB enzyme activity as determined by electrophoresis (n = 203; r = 0.97; slope = 0.90; y-intercept = -4.29) and with commercial immunoassays. We think that this assay will be useful for confirmation of acute myocardial infarction, both in critical-care units and in the clinical laboratory.     

The value of serum CK-MB and myoglobin measurements for assessing perioperative myocardial infarction after cardiac surgery

In 41 patients who underwent coronary bypass surgery, creatine kinase (CK)-MB mass concentration was repeatedly measured in serum during and after the intervention using a new two-site immunoenzymetric assay (IEMA). Serum CK-MB activity was determined with the use of four different techniques: immunoinhibition, immunoinhibition-immunoprecipitation, column chromatography and electrophoresis. Myoglobin (Mb) was also measured in each specimen by radioimmunoassay. In the 33 patients who followed a completely uneventful postoperative course, the cumulated CK-MB release was, on the average, 12.2-fold less than after acute myocardial infarction. The CK-MB peak concentrations using the IEMA were 33 +/- 3 micrograms/l (X +/- SEM) and occurred 6.4 +/- 0.5 h after the intervention was started; CK-MB levels had decreased to 2.9 +/- 0.4 micrograms/l at the end of the first postoperative day. The evolution of the CK-MB concentration was parallel to that of the enzyme activity. The serum Mb maximum concentrations (518 +/- 39 micrograms/l) were reached after 3.3 +/- 0.1 h. The other eight patients developed perioperative myocardial infarction (PMI); in this group, the cumulated CK-MB release was higher, and the serum CK-MB postoperative curves were of three different types. The patients with delayed CK-MB peaks (type I pattern) or sustained elevations (type III) of this isoenzyme also showed increased serum Mb levels at the end of the first postoperative day. The PMI patients with early (10 h) CK-MB elevations (type II) did not demonstrate abnormal serum Mb levels.(ABSTRACT TRUNCATED AT 250 WORDS)     

AMI患者早期CK同工酶亚型的高压电泳图谱分析

目的探讨CK同工酶亚型在AMI胸痛发作后24h内的变化规律,为AMI患者的早期诊断提供依据。方法采用REP全自动高压电泳仪检测AMI胸痛发作后不同时间以及对照组的CK-MM1、CK-MM2、CK-MM3、CK-MB1、CK-MB2等指标并进行荧光扫描,同时在Olympus2700全自动生化分析仪上测定CK、CK-MB的总活性,对所得数据进行恰当的统计分析。结果在AMI患者胸痛发作24h内,CK同工酶亚型有一特殊的变化规律:4~6h内,CK-MB2、CK-MM3开始升高,8~12h达高峰,92%的病人CK-MB2/CK-MB1>1.5,同时91%的病人CK-MM3/CK-MM1>0.5。结论CK同工酶亚型CK-MM3/CK-MM1、CK-MB2/CK-MB1的检测可作为AMI早期诊断的指标。     

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.