Small Extracellular Vesicles/Exosomes Protein Profile

Small Extracellular Vesicles/Exosomes Protein Profile

Exosomes are membrane-bound extracellular vesicles that contain many biomolecules, including proteins, RNA, DNA, etc. Proteins, as a major component of exosomes, are crucial for a deep understanding of exosomal functions and their roles in diseases. The study of exosomal proteomics, which can be conducted using various techniques such as mass spectrometry(MS) and immunological assays, provides researchers with extensive information, including exosome biosynthesis, secretion, cell signal transduction, disease occurrence and development and so on. In addition, the workflow for exosomal proteomic service typically involves critical steps such as exosome extraction, protein isolation, and mass spectrometric analysis. Through these processes, researchers are able to acquire a thorough understanding of exosomal proteins, laying a solid foundation for subsequent analysis and research.

Label-Free Quantitative (LFQ) Protein Profile

Quantitative proteomics is an analytical chemical technique that uses the MS platform to detect the relative content of proteins in samples. Label-free quantitative proteomics is a specific approach that does not rely on isotope labeling for protein quantification. This technology detects the peptide fragments of protein by liquid chromatography-mass spectrometry(LC-MS), analyzes the MS data generated when protein was identified on a large scale, and integrates the detected ion peak intensity to make relative quantification by the integrated area.

Tandem Mass Tag (TMT) Labeled Quantitative Protein Profile

TMT is a relative and absolute quantification technology developed by Thermo Fisher Scientific for the isobaric labeling of peptides in vitro. It utilizes tags with 6, 10, or 16 different isotopes to specifically label the amino groups of peptides. Through analysis by a high-resolution mass spectrometer in tandem mode, it enables the simultaneous comparison of protein expression levels in up to 16 different samples. TMT represents a high-throughput screening technology in quantitative proteomics.

Data-Independent Acquisition (DIA) Quantitative Protein Profile

DIA is emerging as a prominent technique. Unlike the DDA data collection mode, which collects the ions of a limited number of peptides (usually the top 20) with higher response values in each time window of the primary MS for fragmentation analysis, DIA divides the entire full scan range of the mass spectrometer into several windows. All ions in each window are rapidly and cyclically selected, fragmented, and detected, thereby obtaining the complete fragment information of all ions in the sample without omission and difference.

Parallel Reaction Monitoring (PRM) Targeted Protein Profile

PRM is an ion monitoring technology based on high-resolution, high-precision MS, exemplified by the Orbitrap. It starts by utilizing the selection ability of a quadrupole mass analyzer to select the precursor ion of the target peptide with Q1 initially, and then fragments the precursor ion in the collision cell. Finally, the Orbitrap analyzer was used to detect all fragmentation information within the selected precursor ion window in the secondary MS. This enables selective detection of target proteins and peptides (e.g., peptides with post-translational modifications), enabling relative (or absolute) quantification of the target protein/peptide. Thus, it achieves accurate and specific analysis of target proteins/peptides in complex samples.

Macroproteomics

Macroproteomics is a novel technology within the field of proteomics that applies proteomic techniques to the study of microbial communities. It allows for the comprehensive identification of all proteins within a microbial community at a specific point in time, providing valuable functional insights into the role of microorganisms within ecosystems. At the same time, metaproteomics can also link the genetic and functional diversity of microbial communities, and metaproteomic analysis for different environments can capture new functional genes and metabolic pathways, and identify proteins associated with specific costhood. In addition, proteomics combined with metagenomics data can better reveal the taxonomic diversity, functional diversity and biological processes of environmental communities.

Proximity Barcoding Assay (PBA) Monovesicular Membrane Protein Profile

PBA is a technique that utilizes micrometer-sized single-stranded DNA clusters, each of which contains hundreds of unique DNA copy sequences generated through rolling circle amplification. These DNA clusters serve as specific DNA barcodes for individual extracellular vesicles. The surface protein composition of individual extracellular vesicles in a sample is converted into DNA sequence information through antibody-DNA conjugates that bind to them. These conjugates incorporate random coding sequences present in each rolling circle amplification product. After PCR amplification, next-generation sequencing technology is used to decode the DNA sequences containing protein and exosome information, identifying the surface proteome of individual exosomes.

PBA can encode the surface proteins of all exosomes in a sample at high throughput. The detection sensitivity of PBA depends on factors such as protein binding specificity, the scope and depth of sequencing, and the affinity and selectivity of the antibodies used. By comparing extracellular vesicle profiles from different sources, specific combinations of surface proteins can be identified as biomarkers, aiding in the identification and quantification of extracellular vesicles released from specific tissues into the bloodstream in both health and disease contexts.

Services at NextGen Exsome

1.Label-free Quantitative Exosomes Protein Profile

Proteomic analysis is performed using the label-free quantitative method, followed by standard data analysis.

Deliverables:

1)Report in DOCX format