Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2023 Feb:38 Suppl 1:e9431.
doi: 10.1002/rcm.9431. Epub 2023 Jan 3.

A bottom-up proteomics workflow for a system containing multiple organisms

Hongxia Bai  1 , Leonard B Collins  2 , Marcos Rogério André  3   4 , Edward B Breitschwerdt  4 , Taufika Islam Williams  1   2
Affiliations

A bottom-up proteomics workflow for a system containing multiple organisms

Hongxia Bai et al. Rapid Commun Mass Spectrom. 2023 Feb.

Abstract

Rationale: Discovery proteomics has been popularized to be essential in the investigator's biological toolbox. Many biological problems involve the interplay of multiple organisms. Herein, a bottom-up proteomics workflow was developed to study a system containing multiple organisms to promote a thorough understanding of how each interacts with the others.

Methods: A label-free quantification proteomics workflow was developed with nanoscale liquid chromatography coupled to tandem mass spectrometry (nanoLC-MS/MS). This protocol describes a bottom-up proteomics workflow used to study differential protein expression in the context of fleas (Ctenocephalides felis felis) experimentally infected by the bacterium Bartonella henselae, the etiological agent of Cat Scratch Disease (CSD).

Results: Step-by-step instructions are provided for protein extraction, protein cleanup, total protein measurement, nanoLC-MS/MS data acquisition, and data analysis using Proteome Discoverer software. Comprehensive and exhaustive details are included to promote the adoption of this proteomics workflow in other laboratories.

Conclusion: A proteomics protocol is detailed for a system containing multiple proteomes from different taxonomic lineages using CSD (cats bitten by fleas infected with Bartonella henselae) as a model. The operating protocol can be readily applied to other label-free proteomics work involving multiple proteomes from taxonomically distinct organisms.

PubMed Disclaimer

References

REFERENCES

    1. André MR, Neupane P, Lappin M, et al. Using proteomic approaches to unravel the response of Ctenocephalides felis felis to blood feeding and infection with Bartonella henselae. Front Cell Infect Microbiol. 2022;12:828082. doi:10.3389/fcimb.2022.828082
    1. Breitschwerdt EB, Maggi RG, Chomel BB, Lappin MR. Bartonellosis: An emerging infectious disease of zoonotic importance to animals and human beings. J Vet Emerg Crit Care. 2010;20(1):8-30. doi:10.1111/j.1476-4431.2009.00496.x
    1. Laukaitis HJ, Macaluso K. Unpacking the intricacies of rickettsia-vector interactions. Trends Parasitol. 2021;37(8):734-746. doi:10.1016/j.pt.2021.05.008
    1. Contreras M, Villar M, Artigas-Jerónimo S, Kornieieva L, Mуtrofanov S, de la Fuente J. A reverse vaccinology approach to the identification and characterization of Ctenocephalides felis candidate protective antigens for the control of cat flea infestations. Parasit Vectors. 2018;11(1):43. doi:10.1186/s13071-018-2618-x
    1. Wiśniewski J, Zougman A, Nagaraj N, Mann M. Universal sample preparation method for proteome analysis. Nat Methods. 2009;6(5):359-362. doi:10.1038/nmeth.1322

LinkOut - more resources