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A Mass Spectrometry-Based Proteomics Approach for Global and High-Confidence Protein R-Methylation Analysis
Published on: April 28, 2022
Specimen Inactivation Methods for Proteomics─Comparisons of Irradiation, Chemical, and Heat Treatments on Downstream
Brooke N Genovese1,2, Nistara Randhawa2, Gabriela Grigorean3
1One Health Institute, School of Veterinary Medicine, University of California, Davis, California 95616, United States.
Abstract:
Multiomic techniques, including proteomics, can provide novel insights for both pathogen detection and assessment of host responses to infection. Numerous studies have described the efficacy of heat inactivation, γ irradiation, or treatment with guanidinium-based chaotropic salts (e.g., TRIzol) for pathogen inactivation of biological samples to ensure biosafety and biosecurity. However, the effect of these methods on the greater serum proteome is less studied. Here, we sought to comprehensively measure the effects of four routinely used pathogen inactivation methods on the serum proteome of Rhesus macaques by characterizing the serum proteome pre-and-post inactivation treatment. Using the R-package MS-DAP for statistical benchmarking, our analysis revealed distinct changes in total peptide/protein detection and individual protein abundances (e.g., ALB, APOA1, and CRP) across inactivation methods and compared to their untreated controls. Specifically, we observed improved quantitative reproducibility in γ-irradiated samples across biological, technical, and experimental replicates compared to chemical inactivation and different heat combinations. These findings represent the first direct, experimental comparisons of effective pathogen inactivation methods on the serologic profiles of nonhuman primates and provide useful criteria for evaluating methods for biological specimen inactivation prior to proteomic analysis.
Insights
Pathogen inactivation methods like gamma irradiation or heat treatment significantly alter serum proteome profiles in Rhesus macaques. Gamma irradiation offered better quantitative reproducibility for proteomic analysis compared to other methods.
Area of Science:
- Proteomics
- Infectious Disease Research
- Biosafety
Background:
- Multiomic techniques, including proteomics, offer insights into pathogen detection and host responses.
- Pathogen inactivation is crucial for biosafety but its impact on the serum proteome is understudied.
Purpose of the Study:
- To comprehensively measure the effects of four common pathogen inactivation methods on the Rhesus macaque serum proteome.
- To compare these effects pre- and post-treatment using quantitative proteomic analysis.
Main Methods:
- Serum samples from Rhesus macaques were subjected to four different pathogen inactivation methods.
- Proteomic analysis was performed pre- and post-treatment.
- The R-package MS-DAP was used for statistical benchmarking and analysis.
Main Results:
- Distinct changes in total peptide/protein detection and individual protein abundances (e.g., ALB, APOA1, CRP) were observed across methods.
- Gamma irradiation demonstrated improved quantitative reproducibility compared to chemical and heat inactivation methods.
- Significant alterations in the serum proteome were noted relative to untreated controls.
Conclusions:
- Pathogen inactivation methods significantly impact serum proteomic profiles in nonhuman primates.
- Gamma irradiation appears to be a more reproducible method for proteomic analysis compared to chemical or heat treatments.
- These findings provide criteria for selecting appropriate inactivation methods for proteomic studies.
