Department of Molecular Biology, University of Salzburg, Salzburg, Austria.
Sci Rep. 2017 Mar 20;7:44750. doi: 10.1038/srep44750.
Low endotoxin recovery (LER) is a recently discovered phenomenon describing the inability of limulus amebocyte lysate (LAL)-based assays to detect lipopolysaccharide (LPS) because of a "masking effect" caused by chelators or detergents commonly used in buffer formulations for medical products and recombinant proteins. This study investigates the masking capacities of different buffer formulations and whether masked endotoxin is biologically active. We show that both naturally occurring endotoxin as well as control standard endotoxin can be affected by LER. Furthermore, whereas masked endotoxin cannot be detected in Factor C based assays, it is still detectable in a cell-based TLR4-NF-κB-luciferase reporter gene assay. Moreover, in primary human monocytes, masked LPS induces the expression of pro-inflammatory cytokines and surface activation markers even at very low concentrations. We therefore conclude that masked LPS is a potent trigger of immune responses, which emphasizes the potential danger of masked LPS, as it may pose a health threat in pharmaceutical products or compromise experimental results.
低内毒素回收率（LER）是一种最近发现的现象，描述了基于鲎变形细胞溶解物（LAL）的检测方法由于螯合剂或清洁剂的“掩蔽作用”而无法检测脂多糖（LPS），这些物质通常用于医疗产品和重组蛋白的缓冲液配方中。本研究调查了不同缓冲液配方的掩蔽能力，以及掩蔽内毒素是否具有生物活性。我们表明，天然内毒素和对照标准内毒素都可能受到 LER 的影响。此外，虽然基于因子 C 的检测方法无法检测到掩蔽内毒素，但在基于 TLR4-NF-κB-荧光素酶报告基因的细胞检测方法中仍可检测到。此外，在原代人单核细胞中，即使在非常低的浓度下，掩蔽 LPS 也能诱导促炎细胞因子和表面激活标志物的表达。因此，我们得出结论，掩蔽 LPS 是免疫反应的有效触发物，这强调了掩蔽 LPS 的潜在危险，因为它可能在药物产品中构成健康威胁，或影响实验结果。
