Cryo-OrbiSIMS for 3D molecular imaging of a bacterial biofilm in its native state

We developed a method for analysis and imaging of biological samples in their native state, by combining a cryo-OrbiSIMS instrument with cryogenic sample handling and high-pressure freezing. By using this method, we did analysis and imaging of frozen-hydrated mature <em>Pseudomonas aeruginosa</em> biofilm, which allows the identification and map of quorum sensing signaling molecules, nucleobases and bacterial membrane molecules with high spatial-resolution and high mass-resolution. Some of quorum sensing signaling molecules were further confirmed by MS/MS. By comparing the analysis of frozen-hydrated <em>Pseudomonas aeruginosa</em> biofilm with the freeze-dried one, we dicover that signal intensity of all interesting molecules get enhanced in the frozen-hydrated state. Especially for polar molecules, such as amino acid, it could even achieve 10,000 fold increasing. Here, we provide the original OrbiSIMS data including MS and MS/MS spectra, depth profile and images of frozen-hydrated and freeze-dried <em>Pseudomonas aeruginosa</em> biofilm. The data could be open by using SurfaceLab Version 7.0 (ION-TOF, Germany).本研究结合低温OrbiSIMS（cryo-OrbiSIMS）仪器、低温样品处理系统与高压冷冻技术，开发了一种可对生物样本进行原生状态下分析与成像的方法。利用该方法，我们对冷冻水合状态下的成熟铜绿假单胞菌（Pseudomonas aeruginosa）生物被膜开展了分析与成像，实现了对群体感应信号分子、核碱基以及细菌膜分子的高空间分辨率与高质量分辨率鉴定与定位成像。部分群体感应信号分子经串联质谱（MS/MS）验证得以确认。通过对比冷冻水合与冷冻干燥状态下的铜绿假单胞菌生物被膜分析结果，我们发现目标分子的信号强度在冷冻水合状态下均得到提升，其中极性分子（如氨基酸）的信号强度增幅甚至可达10000倍。本数据集包含原始OrbiSIMS数据，涵盖冷冻水合与冷冻干燥状态下铜绿假单胞菌生物被膜的质谱（MS）、串联质谱（MS/MS）谱图、深度剖面以及成像数据。该数据可通过德国ION-TOF公司的SurfaceLab 7.0版本软件进行读取与查看。