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- Proteomics of fish white muscle and Western blotting to detect putative allergensPublication . Anjos, Liliana; Loukissas, A. Ζ.; Power, DeborahA detailed workflow is provided for preparation from teleost fish white muscle of extracts for proteomics analysis. The protocol generates samples that can be analyzed by SWATH (Sequential Window data independent Acquisition of the Total High-resolution-Mass Spectra), a modern MS-based quantitative label free technology. The main steps for the extraction of three independent protein fractions, (1) soluble sarcoplasmic, (2) soluble myofibrillar, and (3) insoluble material, from fish white muscle are detailed. Coupled to the protein extraction protocol a Western blotting approach is outlined for detection of common fish allergens, in this case β-parvalbumin, in the white muscle sarcoplasmic protein fraction.
- Extracting protein from microalgae (Tetraselmis chuii) for proteome analysisPublication . L, Anjos; Estêvão, J.; Infante, Carlos; Mantecón, Lalia; Power, Deborah MaryMicroalgae have high potential as a resource for sustainable and green protein for food or bioactive molecules. Nonetheless, despite the high protein content of microalgae (40 - 70% dry weight) progress in the characterization of their protein composition remains challenging. This is due to the highly variable chemical composition of microalgae strains and factors such as their rigid thick cell wall, polysaccharide content, protein stability, pH. The method described herein was developed to optimize protein extraction for proteome analysis of microalgae (Tetraselmis chuii) biomass. The effects on protein solubility of solvent type (organic, denaturing, and non-denaturing) combined with three customized microalgae disruption methods were investigated. The proteome targeted high quality protein extracts were for hydro-soluble proteins recovered by cell disruption using bead milling coupled to centrifugation (protein yield approximate to 13%). The developed method is inexpensive, efficient (yielding high-quality protein extracts with a low content of interfering compounds) and from an industrial perspective easily scalable and compatible with other applications. To add value to the end product we additionally propose the use of stabilizing agents to maintain protein solubility during refrigerated storage and a method targeting the fractionation of low molecular weight proteins. An inexpensive easy-to-do 5 step protocol for microalgae protein extracts. A protein extraction method free from dangerous or highly polluting chemicals. Production of high yield aqueous protein extracts suitable for proteomics. (C) 2022 The Authors. Published by Elsevier B.V.
- High pressure processing of European sea bass (Dicentrarchus labrax) fillets and tools for flesh quality and shelf life monitoringPublication . Tsironi, Theofania; Anjos, Liliana; Pinto, Patricia IS; Dimopoulos, George; Santos, Soraia; Santa, Cátia; Manadas, Bruno; Canario, Adelino; Taoukis, Petros; Power, DeborahThe effects of high pressure (HP:600 MPa, 5 min, 25 °C) on European sea bass fillets were investigated using microbiological, physicochemical and sensory indices, and “omics” technologies. HPP led to more than a 5 log(cfu/g) reduction in initial bacterial total viable counts and altered the bacterial microbiome, reducing the proportion of food spoilage genera. Lightness and hardness of the fish flesh significantly increased after HPP and were associated with modified muscle tissue histology, with fibers appearing fused and more compact in comparison to the unprocessed control. Sensory evaluation (based on a lower limit of 5 for overall acceptability scoring) indicated a shelf life of 11 days for untreated control samples and 2 months for the HP-treated fillets. Quantitative SWATH proteomics revealed 281 proteins that had modified levels between control and HP-processed fish flesh. The metagenomics and proteomics provided detailed insight into how the change in HP-processed sea bass fillets is linked to the modifications in the microbiome and proteome.