Kochan Kamila, Peleg Anton Y, Heraud Philip, Wood Bayden R
Centre for Biospectroscopy and School of Chemistry, Monash University;
Infection and Immunity Program, Monash Biomedicine Discovery Institute and Department of Microbiology, Monash University; Department of Infectious Diseases, The Alfred Hospital and Central Clinical School, Monash University.
J Vis Exp. 2020 Sep 15(163). doi: 10.3791/61728.
Atomic Force Microscopy-Infrared Spectroscopy (AFM-IR) is a novel combinatory technique, enabling simultaneous characterization of physical properties and chemical composition of sample with nanoscale resolution. By combining AFM with IR, the spatial resolution limitation of conventional IR is overcome, enabling a resolution of 20-100 nm to be achieved. This opens the door for a broad array of new applications of IR toward probing samples smaller than several micrometers, previously unachievable by means of conventional IR microscopy. AFM-IR is eminently suited for bacterial research, providing both spectral and spatial information at the single cell and intracellular level. The increasing global health concerns and unfavorable future prediction regarding bacterial infections, and especially, rapid development of antimicrobial resistance, has created an urgent need for a research tool capable of phenotypic probing at the single cell and subcellular level. AFM-IR offers the potential to address this need, by enabling detail characterization of chemical composition of a single bacterium. Here, we provide a complete protocol for sample preparation and data acquisition of single spectra and mapping modality, for the application of AFM-IR toward bacterial studies.
原子力显微镜-红外光谱技术（AFM-IR）是一种新型的组合技术，能够以纳米级分辨率同时表征样品的物理性质和化学成分。通过将原子力显微镜与红外光谱相结合，克服了传统红外光谱的空间分辨率限制，实现了20-100纳米的分辨率。这为红外光谱在探测小于几微米的样品方面开辟了一系列新的应用前景，而这是传统红外显微镜以前无法实现的。AFM-IR非常适合细菌研究，可在单细胞和细胞内水平提供光谱和空间信息。全球对细菌感染的健康关注度不断提高，以及对细菌感染未来的不利预测，尤其是抗菌药物耐药性的迅速发展，迫切需要一种能够在单细胞和亚细胞水平进行表型探测的研究工具。AFM-IR通过能够详细表征单个细菌的化学成分，提供了满足这一需求的潜力。在此，我们提供了一个完整的方案，用于样品制备以及单光谱和映射模式的数据采集，以将AFM-IR应用于细菌研究。
