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Counting and Recognizing Single Bacterial Cells by a Lanthanide-Encoding Inductively Coupled Plasma Mass Spectrometric Approach

Liang, Yong, Liu, Qian, Zhou, Yang, Chen, Shi, Yang, Limin, Zhu, Min, Wang, Qiuquan
Analytical chemistry 2019 v.91 no.13 pp. 8341-8349
Escherichia coli, Listeria monocytogenes, Shigella dysenteriae, Staphylococcus aureus, Vibrio parahaemolyticus, antibiotic resistance, bacteria, cell walls, detection limit, mass spectrometry, nanosilver, peptidoglycans, polyclonal antibodies, vancomycin
Counting and recognizing single bacterial cells are crucial to the diagnosis of bacterium-induced disease and study of cell-to-cell variability as well as the related antibiotic resistance mechanism. A higher sensitive and selective method has always been desired for a more accurate single bacterial cell analysis. We report a lanthanide-encoding inductively coupled plasma (ICP) mass spectrometric approach for counting and recognizing single bacterial cells for the first time. When noncanonical alkyne-d-alanine (aDA) was added to five typical bacterial strains of Escherichia coli, Staphylococcus aureus, Listeria monocytogenes, Shigella dysenteriae, and Vibrio parahemolyticus, aDA was metabolically assembled into the peptidoglycan layer-supported bacterial cell wall followed by post-clickable europium-tagging with 1,4,7,10-tetraazacyclododecane-1,4,7-tris-acetic acid-10-azidopropyl ethylacetamide-europium complex (azide-DOTA-Eu). Such Eu-tagged bacterial cells can be deemed as Eu-engineered particles, delivering more than 5 orders of magnitude self-signal-amplification outcome relative to the single bacterial cells themselves when ¹⁵¹/¹⁵³Eu is determined by single particle ICP mass spectrometry (spICPMS). This metabolic assembly of aDA mediated Eu-encoding signal amplification strategy breaks through the detection limit of spICPMS and ensures that we directly count a single bacterial cell. The individual bacterial strains we counted can be simultaneously recognized through their corresponding lanthanide (Ln)-coded polyclonal antibody (Ln = ¹³⁹La, ¹⁴¹Pr, ¹⁴²Nd, ¹⁵²Sm, and ¹⁶⁰Gd, respectively), serving as a specific bacterial identification (Ln-pAb-ID). Moreover, the developed approach was applied to show the different behavior between genetically identical Staphylococcus aureus under the treatments of vancomycin and Ag nanoparticles, demonstrating that such a lanthanide-encoding spICPMS approach provided a new way to discover still ambiguous cell-to-cell variability.