Norharmane matrix enhances detection of endotoxin by MALDI-MS for simultaneous profiling of pathogen, host and vector systems
Norharmane matrix enhances detection of endotoxin by MALDI-MS for simultaneous profiling of pathogen, host and vector systems
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DOI：
10.1093/femspd/ftw097
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发表时间：
2016-11-01
影响因子：
3.3
通讯作者：
Ernst, Robert K.
中科院分区：
文献类型：
--
作者：
Scott, Alison J.;Flinders, Bryn;Ernst, Robert K.
细菌感染过程中新的致病机制的发现需要先进技术的发展。在这里，我们评估了双极性基质去甲肾上腺素(NRM)以改进从感染新城疫杆菌(FN)的宿主和载体组织中检测细菌脂A(内毒素)的方法。我们对NRM进行了评估，以改进正负两种极性的广泛脂类的检测和表征，包括基质辅助激光解吸-电离耦合应用中的类脂A和磷脂。NRM基质将单磷脂A的检测限(LOD)提高到皮克水平，表明LOD比2，5-二羟基苯甲酸提高了10倍，LOD比9-氨基吖啶(9-AA)提高了100倍。随后，改进的脂类A的LOD有助于从受感染的小鼠脾提取液中检测主要离子(m/z 1665)，以及从受感染的变色粉虱中检测到m/z为1637的温度修饰的Fn脂类A。最后，我们同时定位了FN感染的脾内的细菌磷脂特征以及宿主独有的基于肌醇的磷脂(m/z 933)，展示了宿主-病原体相互作用的共图谱。在其他感染模型和内毒素靶向成像实验中扩大使用NRM基质将提高我们对宿主-病原体界面上的脂质相互作用的理解。
The discovery of novel pathogenic mechanisms engaged during bacterial infections requires the evolution of advanced techniques. Here, we evaluate the dual polarity matrix norharmane (NRM) to improve detection of bacterial lipid A (endotoxin), from host and vector tissues infected with Francisella novicida (Fn). We evaluated NRM for improved detection and characterization of a wide range of lipids in both positive and negative polarities, including lipid A and phospholipids across a range of matrix-assisted laser desorption-ionization-coupled applications. NRM matrix improved the limit of detection (LOD) for monophosphoryl lipid A (MPLA) down to picogram level representing a 10-fold improvement of LOD versus 2,5-dihydroxybenzoic acid and 100-fold improvement of LOD versus 9-aminoacridine (9-AA). Improved LOD for lipid A subsequently facilitated detection of the Fn lipid A major ion (m/z 1665) from extracts of infected mouse spleen and the temperature-modified Fn lipid A at m/z 1637 from infected Dermacentor variabilis ticks. Finally, we simultaneously mapped bacterial phospholipid signatures within an Fn-infected spleen along with an exclusively host-derived inositol-based phospholipid (m/z 933) demonstrating coprofiling of the host-pathogen interaction. Expanded use of NRM matrix in other infection models and endotoxin-targeting imaging experiments will improve our understanding of the lipid interactions at the host-pathogen interface.