Citation: NIU Qian, TIAN Ye, WU Ming-Kai, ZHANG Yu-Hao, CHEN Chen, YAN Xiao-Mei. Quantitative Characterization of Purity, Concentration and Size Distribution of Bacterial Membrane Vesicles at Single-Particle Level[J]. Chinese Journal of Analytical Chemistry, ;2021, 49(5): 733-742. doi: 10.19756/j.issn.0253-3820.211054 shu

Quantitative Characterization of Purity, Concentration and Size Distribution of Bacterial Membrane Vesicles at Single-Particle Level

Departmentof Chemical Biology, MOE Key Laboratory of Spectrochemical Analysis & Instrumentation, Key Laboratory for Chemical Biology of Fujian Province, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005, China

Corresponding author: YAN Xiao-Mei, xmyan@xmu.edu.cn
Received Date: 22 January 2021
Revised Date: 23 March 2021

Fund Project: Supported by the National Natural Science Foundation of China (Nos.21934004, 21627811) and the National Fund for Fostering Talents of Basic Science (No.J1310024).

Bacterial membrane vesicles (BMVs) are nanostructures shed naturally by most bacterial species. BMVs carry nucleic acids, proteins, especially virulence factors and immune regulatory factors for the inter- and intra-species communication, showing great potential in the development of vaccines and anticancer drugs. However, the purification and characterization of BMVs remain challenging due to their large heterogeneity, nanoscale particle size, and low contents of cargo molecules. Based on the unique advantages such as the laboratory-built nano-flow cytometer (nFCM) in the sensitive and rapid analysis of individual nanoparticles, the purity, concentration and size distribution of BMV preparations were measured in this work. The experimental results demonstrated that the components in each density layer after density gradient centrifugation could be detected by nFCM to obtain BMVs samples with a high purity of more than 90%. BMVs secreted by Gram-positive bacteria S.aureus and Gram-negative bacteria E.coli O157:H7 were significantly different in density, which mainly presented in the second (ρ=1.06 g/mL) and fourth (ρ=1.10 g/mL) fractions of the Optiprep density gradient layer respectively. The median particle sizes of BMVs derived from Gram-positive (S.aureus) and Gram-negative (E.coli O157:H7) bacteria were 64.9 nm and 83.5 nm, respectively. In conclusion, the study presented here reported the development of an efficient method to characterize the purity, concentration, and size distribution of BMVs at the single-particle level. nFCM was expected to provide powerful technical support for the biological function study of BMVs.
- Bacterial membrane vesicle,
- Nano-flow cytometer,
- Density gradient centrifugation,
- Purity,
- Concentration,
- Size distribution
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