Landt Y, Vaidya H C, Porter S E, Whalen K, McClellan A, Amyx C, Parvin C A, Kessler G, Nahm M H, Dietzler D N
Department of Pathology, Washington University School of Medicine, St. Louis, MO 63110.
Clin Chem. 1988 Mar;34(3):575-81.
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.
这种用于肌酸激酶MB同工酶（EC 2.7.3.2）的半自动比色测定法基于一种针对该同工酶的单克隆抗体（“Conan-MB”），是对先前发表方法的改进（Vaidya等人，《临床化学》1986年；32:657 - 63）。将涂有2至3微克抗体的0.64厘米珠子与100微升血清和10微升0.2摩尔/升β-巯基乙醇在室温下孵育1小时，以提取CK-MB。珠子用去离子水洗涤，然后在37℃下与CK底物孵育45分钟。加入含有反式-1,2-二氨基环己烷-N,N,N',N'-四乙酸、对碘硝基四氮唑紫和黄递酶的溶液，并在492纳米处测量产生的有色产物。标准曲线在每升200 U的CK-MB范围内呈线性，分析回收率为97 - 113%。低质量控制品（9.7 U/L）和高质量控制品（50.7 U/L）的总分析变异系数分别为14.1%（n = 1878）和11.6%（n = 1842）。CK-MB活性与通过两点质量免疫测定法测得的CK-MB相关性良好（r = 0.978，n = 226），并且99.4%的CK-MB大于或等于12 U/L的样本（n = 347）通过琼脂糖电泳得到验证。
