Robust optimization of SWATH-MS workflow for human blood serum proteome analysis using a quality by design approach

Abstract Background It is not enough to optimize proteomics assays. It is critical those assays are robust to operating conditions. Without robust assays, proteomic biomarkers are unlikely to translate readily into the clinic. This study outlines a structured approach to the identification of a robust operating window for proteomics assays and applies that method to Sequential Window Acquisition of all Theoretical Spectra Mass Spectroscopy (SWATH-MS). Methods We used a sequential quality by design approach exploiting a fractional screening design to first identify critical SWATH-MS parameters, then using response surface methods to identify a robust operating window with good reproducibility, before validating those settings in a separate validation study. Results The screening experiment identified two critical SWATH-MS parameters. We modelled the number of proteins and reproducibility as a function of those parameters identifying an operating window permitting robust maximization of the number of proteins quantified in human serum. In a separate validation study, these settings were shown to give good proteome-wide coverage and high quantification reproducibility. Conclusions Using design of experiments permits identification of a robust operating window for SWATH-MS. The method gives a good understanding of proteomics assays and greater data-driven confidence in SWATH-MS performance.

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Robust Optimization of SWATH-MS workflow for human blood serum proteome analysis using a Quality by Design approach

10.21203/rs.3.rs-347514/v1 ◽

2021 ◽

Author(s):

Edith Serrano-Blesa ◽

Andrew Porter ◽

Dennis William Lendrem ◽

Costantino Pitzalis ◽

Anne Barton ◽

...

Keyword(s):

Validation Study ◽

Quality By Design ◽

Proteome Analysis ◽

Operating Conditions ◽

Design Approach ◽

Screening Design ◽

Study Results ◽

Quality By Design Approach ◽

Operating Window ◽

Structured Approach

Abstract Background: It is not enough to optimize proteomics assays. It is critical those assays are robust to operating conditions. Without robust assays, proteomic biomarkers are unlikely to translate readily into the clinic. This study outlines a structured approach to the identification of a robust operating window for proteomics assays and applies that method to Sequential Window Acquisition of all Theoretical Spectra Mass Spectroscopy (SWATH-MS). Methods: We used a sequential Quality by Design approach exploiting a fractional screening design to first identify critical SWATH-MS parameters, then using response surface methods to identify a robust operating window with good reproducibility, before validating those settings in a separate validation study. Results: The screening experiment identified two critical SWATH-MS parameters. We modelled the number of proteins and reproducibility as a function of those parameters identifying an operating window permitting robust maximization of the number of proteins quantified in human serum. In a separate validation study, these settings were shown to give good proteome-wide coverage and high quantification reproducibility. Conclusions: Using design of experiments permits identification of a robust operating window for SWATH-MS. The method gives a good understanding of proteomics assays and greater data-driven confidence in SWATH-MS performance.

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Resveratrol Nanocrystal Incorporated into Mesoporous Material: Rational Design and Screening through Quality-by-Design Approach

Nanomaterials ◽

10.3390/nano12020214 ◽

2022 ◽

Vol 12 (2) ◽

pp. 214

Author(s):

Ahmad Ainurofiq ◽

Yuniawan Hidayat ◽

Eva Y. P. Lestari ◽

Mayasri M. W. Kumalasari ◽

Syaiful Choiri

Keyword(s):

Particle Size ◽

Rational Design ◽

Quality By Design ◽

Drug Loading ◽

Fractional Factorial ◽

Design Approach ◽

Grape Seeds ◽

Screening Process ◽

Screening Design ◽

Quality By Design Approach

Bioflavonoids from grape seeds feature powerful antioxidant and immunostimulant activities, but they present problems related to solubility and bioavailability. Nanocrystal (NC) incorporated into a mesoporous carrier is a promising strategy to address these issues. However, the preparation of this formulation involves the selection of factors affecting its critical quality attributes. Hence, this study aimed to develop an NC formulation incorporating resveratrol into a soluble mesoporous carrier based on rational screening design using a systematic and continuous development process, the quality-by-design paradigm. A mesoporous soluble carrier was prepared by spray-drying mannitol and ammonium carbonate. The NC was obtained by introducing the evaporated solvent containing a drug/polymer/surfactant and mesoporous carrier to the medium. A 26−2 fractional factorial design (FFD) approach was carried out in the screening process to understand the main effect factors. The type and concentration of polymer and surfactant, resveratrol loading, and solvent were determined on the NC characteristics. The results indicated that drug loading, particle size, and solubility were mainly affected by RSV loading, PEG concentration, and Kolliphor EL concentration. The polymer contributed dominantly to reducing the particle size and enhancing solubility in this screening design. The presence of surfactants in this system made it possible to prolong the supersaturation process. According to the 26−2 FFD, the factors selected to be further developed using a statistical technique according to the quality-by design-approach, Box Behnken Design, were Kolliphor EL, PEG400, and RSV loading.

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