Myoglobin (MG) is a biomarker for heart muscle injury, making it a potential target protein for early detection of myocardial infarction. Elevated myoglobin levels alone have low specificity for acute myocardial infarction (AMI) but in combination with cardiac troponin T have been considered highly efficient diagnostic biomarkers. Myoglobin is a monomeric heme protein with a molecular weight of 17 kDa that is found in skeletal and cardiac tissue as an intracellular storage unit of oxygen. MG consists of eight α-helices connected by loops and a heme group responsible for oxygen-binding. Monoclonal antibodies are widely used analytical tools in biomedical research and have been employed for immunoanalytical detection of MG. However, the epitope(s) recognized by MG antibodies have been hitherto unknown. Precise molecular identification of the epitope(s) recognized by antibodies is of key importance for the development of MG as a diagnostic biomarker. The epitope of a monoclonal MG antibody was identified by proteolytic epitope extraction mass spectrometry in combination with surface plasmon resonance (SPR) biosensor analysis. The MG antibody was immobilized both on an affinity microcolumn and a gold SPR chip. The SPR kinetic analysis provided an affinity-binding constant KD of 270 nM for MG. Binding of a tryptic peptide mixture followed by elution of the epitope from the SPR-MS affinity interface by mild acidification provided a single-epitope peptide located at the C-terminus [146-153] [YKELGFQG] of MG. The specificity and affinity of the epitope were ascertained by synthesis and affinity-mass spectrometric characterization of the epitope peptide.

中文翻译：

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通过 SPR-生物传感器质谱法测定心肌梗塞标志物肌红蛋白的抗体表位和亲和力。

肌红蛋白 (MG) 是心肌损伤的生物标志物，使其成为早期检测心肌梗死的潜在靶蛋白。单独升高的肌红蛋白水平对急性心肌梗死 (AMI) 的特异性较低，但与心肌肌钙蛋白 T 结合被认为是高效的诊断生物标志物。肌红蛋白是一种单体血红素蛋白，分子量为 17 kDa，存在于骨骼和心脏组织中，是细胞内氧的储存单位。MG 由八个通过环连接的 α-螺旋和一个负责氧结合的血红素组组成。单克隆抗体是生物医学研究中广泛使用的分析工具，已被用于 MG 的免疫分析检测。然而，MG抗体识别的表位迄今尚不清楚。对抗体识别的表位进行精确的分子鉴定对于 MG 作为诊断生物标志物的发展至关重要。通过蛋白水解表位提取质谱结合表面等离子共振 (SPR) 生物传感器分析，鉴定了单克隆 MG 抗体的表位。MG 抗体固定在亲和微柱和金 SPR 芯片上。SPR 动力学分析为 MG 提供了 270 nM 的亲和力结合常数 KD。结合胰蛋白酶肽混合物，然后通过温和酸化从 SPR-MS 亲和界面洗脱表位，提供位于 MG C 端 [146-153] [YKELGFQG] 的单表位肽。