Allergy and Clinical Immunology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, 1095 Willowdale Road, Morgantown, WV 26505, USA.
J Immunol Methods. 2013 Nov 29;397(1-2):66-70. doi: 10.1016/j.jim.2013.08.015. Epub 2013 Sep 4.
Diisocyanates (dNCOs) are highly reactive low molecular weight chemicals commonly used in the manufacturing industry. Occupational exposures to dNCOs have been shown to elicit allergic sensitization and occupational asthma. Among the most commonly used dNCOs in industry are the aromatic dNCOs, toluene diisocyanate (TDI) and methylene diphenyl diisocyanate (MDI). This study aimed to develop enzyme linked immunosorbent assays (ELISA) utilizing aromatic dNCO-specific monoclonal antibodies (mAbs) for the detection of aromatic dNCO adducts. Two sandwich ELISAs were developed. The first sandwich ELISA utilized mAb 60G2 along with an anti-human serum albumin (HSA) polyclonal antibody. This assay detected MDI-, 2,4- and 2,6-TDI-HSA adducts with limits of detection (LOD) of 2.67, <0.10, and 1.70 ng/mL, respectively. When spiked into human serum, the LOD of this ELISA increased to 34.37, 7.64 and 24.06 ng/mL, respectively. The second ELISA utilized mAbs 62G5 and 60G2 for capture and detection. This assay was capable of detecting 2,4- and 2,6-TDI-HSA adducts with LODs of <4.90 and 26.92 ng/mL, respectively, and when spiked in human serum, <4.90 and 95.93 ng/mL, respectively. This 62G5-60G2 sandwich assay was also able to detect dNCO adducted transferrin, hemoglobin, keratin and actin, but with less sensitivity than dNCO-HSA. The results of this study demonstrate potential application of these ELISAs in the identification and characterization of aromatic dNCO adducts as well as in biomonitoring occupational and environmental dNCO exposures.
二异氰酸酯（dNCOs）是一种高度反应性的低分子量化学物质，通常用于制造业。职业性接触 dNCOs 已被证明会引起过敏致敏和职业性哮喘。在工业中最常用的芳香族二异氰酸酯（dNCOs）是甲苯二异氰酸酯（TDI）和二苯基甲烷二异氰酸酯（MDI）。本研究旨在开发利用芳香族 dNCO 特异性单克隆抗体（mAbs）检测芳香族 dNCO 加合物的酶联免疫吸附测定（ELISA）。开发了两种夹心 ELISA。第一种夹心 ELISA 利用 mAb 60G2 与抗人血清白蛋白（HSA）多克隆抗体一起使用。该测定法检测 MDI、2,4-和 2,6-TDI-HSA 加合物的检测限（LOD）分别为 2.67、<0.10 和 1.70 ng/mL。当加入人血清中时，该 ELISA 的 LOD 分别增加到 34.37、7.64 和 24.06 ng/mL。第二种 ELISA 利用 mAbs 62G5 和 60G2 进行捕获和检测。该测定法能够检测 2,4-和 2,6-TDI-HSA 加合物，LOD 分别为<4.90 和 26.92 ng/mL，当加入人血清中时，LOD 分别为<4.90 和 95.93 ng/mL。这种 62G5-60G2 夹心测定法还能够检测 dNCO 结合转铁蛋白、血红蛋白、角蛋白和肌动蛋白，但敏感性低于 dNCO-HSA。本研究的结果表明，这些 ELISA 具有在识别和表征芳香族 dNCO 加合物以及生物监测职业和环境 dNCO 暴露方面的潜在应用。
