Healing Foundation Cleft and Craniofacial Clinical Research Centre, University of Manchester, Manchester M13 9WL, United Kingdom.
Department of Nutritional Science, Faculty of Health and Medical Sciences, University of Surrey, Guildford GU2 7XH, United Kingdom; and.
J Lipid Res. 2013 Aug;54(8):2273-2281. doi: 10.1194/jlr.D037432. Epub 2013 May 20.
HDL subclasses detection, in cardiovascular risk, has been limited due to the time-consuming nature of current techniques. We have developed a time-saving and reliable separation of the principal HDL subclasses employing iodixanol density gradient ultracentrifugation (IxDGUC) combined with digital photography. HDL subclasses were separated in 2.5 h from prestained plasma on a three-step iodixanol gradient. HDL subclass profiles were generated by digital photography and gel scan software. Plasma samples (n = 46) were used to optimize the gradient for the resolution of HDL heterogeneity and to compare profiles generated by IxDGUC with gradient gel electrophoresis (GGE); further characterization from participants (n = 548) with a range of lipid profiles was also performed. HDL subclass profiles generated by IxDGUC were comparable to those separated by GGE as indicated by a significant association between areas under the curve for both HDL2 and HDL3 (HDL2, r = 0.896, P < 0.01; HDL3, r = 0.894, P < 0.01). The method was highly reproducible, with intra- and interassay coefficient of variation percentage < 5 for percentage area under the curve HDL2 and HDL3, and < 1% for peak Rf and peak density. The method provides time-saving and cost-effective detection and preparation of the principal HDL subclasses.
由于当前技术的耗时性质，在心血管风险中对 HDL 亚类的检测受到限制。我们开发了一种节省时间且可靠的分离主要 HDL 亚类的方法，该方法结合使用了碘克沙醇密度梯度超速离心（IxDGUC）和数字摄影。在三步碘克沙醇梯度上，可在 2.5 小时内从预染色血浆中分离 HDL 亚类。通过数字摄影和凝胶扫描软件生成 HDL 亚类图谱。使用血浆样本（n = 46）优化梯度以分辨 HDL 异质性，并比较 IxDGUC 与梯度凝胶电泳（GGE）生成的图谱；还对具有不同脂质谱的参与者（n = 548）进行了进一步的特征描述。IxDGUC 生成的 HDL 亚类图谱与 GGE 分离的图谱相当，这表明两种 HDL2 和 HDL3 的曲线下面积之间存在显著关联（HDL2，r = 0.896，P < 0.01；HDL3，r = 0.894，P < 0.01）。该方法具有高度可重复性，HDL2 和 HDL3 曲线下面积百分比的日内和日间变异系数百分比<5，峰 Rf 和峰密度的变异系数百分比<1%。该方法提供了一种节省时间且经济有效的主要 HDL 亚类的检测和制备方法。
