Miyashita Kazuya, Nakajima Katsuyuki, Fukamachi Isamu, Muraba Yuji, Koga Takafumi, Shimomura Yohnosuke, Machida Tetsuyo, Murakami Masami, Kobayashi Junji
Immuno-Biological Laboratories Co., Ltd., Fujioka, Gunma, Japan.
Department of General Medicine, Kanazawa Medical University, Ishikawa, Japan; Hidaka Hospital, Takasaki, Japan.
J Lipid Res. 2017 Aug;58(8):1591-1597. doi: 10.1194/jlr.M075432. Epub 2017 Jun 20.
There is no established method for measuring human hepatic triglyceride (TG) lipase (HTGL) concentration in serum. In this study, we developed new monoclonal Abs (MoAbs) (9A1 mouse MoAb and 141A1 rat MoAb) that react with HTGL both in serum and in postheparin plasma (PHP) and established a novel ELISA system for measuring serum HTGL and PHP-HTGL concentrations. To confirm the specificity of MoAbs, we performed immunoprecipitation-immunoblotting analysis. Both 9A1 mouse MoAb and 141A1 rat MoAb were able to immunoprecipitate not only recombinant HTGL and PHP-HTGL but also serum HTGL, demonstrating that HTGL exists in serum obtained without heparin injection. This method yielded intra- and interassay coefficients of variation of <6% and showed no cross-reactivity with LPL or endothelial lipase. In clinical analysis on 42 male subjects with coronary artery disease, there were strong positive correlations of serum HTGL concentration to PHP-HTGL concentration ( = 0.727, < 0.01). Serum HTGL concentrations showed positive correlations to serum TGs ( = 0.314, < 0.05) and alanine aminotransferase ( = 0.406, < 0.01), and tendencies toward positive correlations to LDL cholesterol, small dense LDL, and γGTP. These results suggest that this new ELISA method for measuring serum HTGL is applicable in daily clinical practice.
目前尚无测定血清中人类肝甘油三酯(TG)脂肪酶(HTGL)浓度的确立方法。在本研究中,我们开发了新的单克隆抗体(MoAbs)(9A1小鼠MoAb和141A1大鼠MoAb),其可与血清和肝素后血浆(PHP)中的HTGL发生反应,并建立了一种新型酶联免疫吸附测定(ELISA)系统来测量血清HTGL和PHP-HTGL浓度。为了确认MoAbs的特异性,我们进行了免疫沉淀-免疫印迹分析。9A1小鼠MoAb和141A1大鼠MoAb不仅能够免疫沉淀重组HTGL和PHP-HTGL,还能免疫沉淀血清HTGL,这表明在未注射肝素获得的血清中存在HTGL。该方法的批内和批间变异系数均<6%,且与脂蛋白脂肪酶(LPL)或内皮脂肪酶无交叉反应。在对42名男性冠心病患者的临床分析中,血清HTGL浓度与PHP-HTGL浓度呈强正相关( = 0.727, < 0.01)。血清HTGL浓度与血清TGs呈正相关( = 0.314, < 0.05),与丙氨酸转氨酶呈正相关( = 0.406, < 0.01),与低密度脂蛋白胆固醇、小而密低密度脂蛋白和γ-谷氨酰转肽酶(γGTP)呈正相关趋势。这些结果表明,这种用于测量血清HTGL的新型ELISA方法适用于日常临床实践。