{"database":"Pride","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Msf":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/MtoZ-ZJL0415202503.msf"],"Txt":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/checksum.txt"],"Raw":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/M1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/M3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/Y2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/Y1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/Y3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/M2.raw"],"Fasta":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/Aloe_vera-20250812.fasta"],"Other":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD069742/MtoZ-ZJL0415202503.pdResult"]},"type":"primary"},"statusCode":"OK","statusCodeValue":200}],"scores":null,"additional":{"labhead_mail":["kwbme@rit.edu"],"submitter":["Stefan Schulze"],"technology_type":["Data-dependent acquisition","Mass Spectrometry","Bottom-up proteomics"],"software":["Not available"],"submitter_keywords":["Oxidative stress","Wound healing","Inflammation resolution","Myofibroblast differentiation","Regenerative skin therapy."],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD069742"],"tissue":["Plant Cell"],"sample_protocol":["To investigate molecular differences in vesicle cargo, label-free data-dependent acquisition (DDA)-based proteomic profiling was carried out on Av-EVs derived from both mature and young Aloe vera leaves. Each group consisted of three biological replicates (n=3), where the mature EV samples were isolated from three independent leaves, and the young EV samples were obtained from three separate potted plants. Following ultracentrifugation, EV pellets were directly resuspended in Pierce™ RIPA lysis buffer (Thermo Scientific, USA, 89901), and protein concentration was determined using the PierceTM BCA protein assay kit (Thermo Scientific, USA, 23225).   Samples were submitted to MtoZ Biolabs (USA) for protein extraction, LC-MS/MS analysis, and downstream bioinformatics employing a transcriptome-informed Aloe vera database (~77,000 protein sequences). In their facilities, proteins were reduced with 10 mM dithiothreitol at 56 °C for 1 h, alkylated with 55 mM iodoacetamide at RT in the dark for 1 h, and digested overnight with sequencing-grade trypsin (Promega) in 50 mM NH₄HCO₃ at 37 °C, using the previously described SP3 workflow. Peptides were desalted using C18 columns, vacuum-dried at 45 °C, and resuspended in 0.1% formic acid. Peptide separation was carried out using an Easy-nLC 1200 system coupled to an Orbitrap Exploris 480 (Thermo Fisher Scientific) with a 66-minute gradient on a C18 column (150 μm i.d. × 170 mm, 1.9 μm, 100 Å). Mass spectrometry was performed in positive ion mode using a top-20 data-dependent acquisition method with a full scan range of m/z 400–1200 at 60,000 resolutions. Peptide fragmentation was achieved using high-energy collision dissociation at a normalized collision energy of 30%."],"repository":["Pride"],"quantification_method":[""],"modification":[""],"data_protocol":["Raw MS/MS data were analyzed against Aloe vera specific database that was generated based on the transcriptomics data described by Choudhri et al. Briefly, unigenes were downloaded from Bioproject PRJNA359629, and coding sequences were predicted using TransDecoder implemented in Galaxy, resulting in 76,958 protein sequences. Search parameters included up to two missed cleavages, fixed carbamidomethylation of cysteines, and variable modifications including methionine oxidation and N-terminal acetylation. A 1% false discovery rate was applied at both peptide and protein levels. Quantitative data were median-normalized and missing values imputed using the K-nearest neighbor (KNN) algorithm. Bioinformatics analyses included differential protein expression, principal component analysis (PCA), hierarchical clustering, gene ontology (GO), Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment, and protein–protein interaction (PPI) network mapping."],"omics_type":["Proteomics"],"labhead":["Karin Wuertz-Kozak"],"instrument_platform":[""],"labhead_affiliation":["Department of Biomedical Engineering, Rochester Institute of Technology, Rochester, USA"],"submission_type":["PARTIAL"],"species":["Aloe Vera"],"submitter_mail":["sschulze@sas.upenn.edu"],"publication":["42037773 Ceballos-Santa CM, Sanchez-Ortiz I, Gaborski TR, Wuertz-Kozak K. A Step-by-Step Protocol for the Isolation of &lt;i&gt;Aloe vera&lt;/i&gt;-Derived Extracellular Vesicles via Manual and Shear-Force Homogenization. Bio Protoc. 2026 16(8):e5664 10.21769/bioprotoc.5664"],"submitter_affiliation":["Rochester Institute of Technology"],"submitter_country":["United States"],"pubmed_abstract":["<i>Aloe vera</i> has long been used for its diverse pharmacological properties, motivating continued interest in isolating and preserving the bioactive molecules responsible for its therapeutic potential. More recently, <i>Aloe vera</i>-derived extracellular vesicles (Av-EVs) have emerged as nanoscale, cell-free carriers capable of retaining and delivering these properties, making them attractive for various biomaterials, nanomedicine, and regenerative medicine applications. Multiple techniques are available for extracellular vesicle isolation. These include ultracentrifugation, polymer-based precipitation, size-exclusion chromatography, immunoaffinity capture, ultrafiltration, density gradient separation, and emerging microfluidic platforms. Each method presents distinct trade-offs in purity, yield, scalability, and downstream compatibility. Despite this diversity, standardized workflows tailored to Av-EV isolation remain limited, and the influence of homogenization-induced shear forces and plant maturity on vesicle recovery and characterization has not been systematically addressed. Here, we present a reproducible protocol for isolating Av-EVs from <i>Aloe vera</i> gel employing two distinct homogenization strategies: manual, no-shear force (NB EVs), and blender-based shear-force homogenization (B EVs). The workflow covers gel preparation, serial centrifugation for debris removal, ultracentrifugation as the gold standard for vesicle enrichment, and final sterile filtration. This protocol enables consistent recovery of Av-EVs suitable for physicochemical characterization and functional analyses. It is simple and relies on commonly available laboratory equipment, facilitating broad adoption by ultracentrifugation users and offering adaptability to diverse research projects involving purified <i>Aloe vera</i> gel and Av-EVs, including studies focused on wound healing, fibrotic scarring, and regenerative processes, where coordinated antioxidant, anti-inflammatory, antimicrobial, immunomodulatory, and moisturizing responses are of interest. Key features • This protocol allows direct comparison of vesicle yield, size distribution, and protein content across extraction methods. • This protocol yields ~1.4-2.0 × 10<sup>10</sup> particles/mL per mature leaf for a total of ~8 × 10<sup>12</sup> particles per leaf. • This protocol yields ~1.2-2.8 × 10<sup>10</sup> particles/mL per young leaf for a total of ~2.8 × 10<sup>12</sup> per leaf. • EVs from mature <i>Aloe</i> leaves yield protein concentrations of ~160-447 μg/mL, corresponding to ~3,840-10,728 μg of protein per leaf."],"pubmed_title":["A Step-by-Step Protocol for the Isolation of &lt;i&gt;Aloe vera&lt;/i&gt;-Derived Extracellular Vesicles via Manual and Shear-Force Homogenization."],"pubmed_authors":["Ceballos-Santa Camila M CM, Sanchez-Ortiz Isabella I, Gaborski Thomas R TR, Wuertz-Kozak Karin K"],"additional_accession":[]},"is_claimable":false,"name":"Targeting Fibrotic Scars with Extracellular Vesicles Extracted from Mature Aloe vera: Enhanced Antioxidant, Anti-inflammatory, and Antifibrotic Activity through M2 Macrophage Polarization and Myofibroblasts Inhibition.","description":"This study aimed to isolate and characterize Av-EVs and evaluate their antioxidant, anti-inflammatory, and antifibrotic properties in vitro, focusing on the impact of extraction method and plant 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