Untargeted Exosome Metabolomics Service

Q: Q1：What is the difference between untargeted and targeted exosome metabolomics?

A: Untargeted exosome metabolomics aims to detect and identify as many metabolites as possible in exosomes without prior knowledge of specific targets, providing a comprehensive overview of the metabolic profile. In contrast, targeted metabolomics focuses on quantifying a predefined set of known metabolites with high specificity and sensitivity, which is often used for validating findings from non-targeted studies.

Q: Q2: How pure do exosome samples need to be for untargeted metabolomics analysis?

A: High-purity exosome samples are crucial to avoid interference from other cellular components or contaminants. Ideally, exosomes should be isolated using methods that minimize co-isolation of proteins, lipoproteins, or cell debris (e.g., ultracentrifugation combined with size-exclusion chromatography). We recommend verifying exosome purity using markers like CD63, CD81, or TSG101 via Western blot or flow cytometry before submission.

Q: Q3: Can untargeted exosome metabolomics detect low-abundance metabolites?

A: Yes, our platform utilizes Orbitrap-based ultra-high-resolution mass spectrometry coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS), a technology renowned for its exceptional sensitivity. This enables us to detect low-abundance metabolites in exosomes, such as certain signaling lipids or rare amino acid derivatives, thereby ensuring comprehensive metabolic profiling.

Q: Q4: Is there a minimum number of samples required for an untargeted exosome metabolomics study?

A: To ensure robust statistical analysis, we recommend a minimum of 5-6 biological replicates per experimental group. This helps account for biological variability and increases the reliability of differential metabolite findings. For complex experimental designs, we can advise on sample size based on your specific research goals.

Q: Q5: How are metabolites identified and annotated in untargeted exosome metabolomics?

A: Metabolites are identified by matching their mass-to-charge ratio (m/z), retention time, and fragmentation patterns against high-quality databases such as METLIN, HMDB, and LipidMaps. We use strict criteria to ensure confident annotations, and provide detailed reports with metabolite names, chemical structures, and confidence scores.

Q: Q6: Can you analyze exosomes from frozen samples that have been stored for a long time?

A: While fresh samples are ideal, we can analyze exosomes from frozen samples stored at -80°C for up to 2-3 years, provided they have not been subjected to multiple freeze-thaw cycles. Repeated thawing can degrade metabolites or damage exosomes, so we recommend aliquoting samples before storage to minimize handling. For samples stored longer than 3 years, we suggest a preliminary quality check to assess metabolite integrity.

Exosome untargeted metabolomics is a professional service that comprehensively analyzes the metabolite composition in exosomes. Creative Biostructure conducts unbiased metabolite analysis on exosomes from different samples through advanced detection technologies. Our advanced technical platform can capture information on a wide range of small molecule metabolites in exosomes, providing comprehensive and accurate data support for customers in life science research, disease mechanism exploration, biomarker discovery, etc., and helping customers explore the biological significance behind exosome metabolites.

Why is it Necessary to Measure Metabolites in Exosomes?

As nanoscale vesicles secreted by cells, exosomes are widely present in various biological fluids and cell culture media. They carry a large number of biologically active molecules, among which metabolites are an important component. These metabolites are involved in various physiological processes such as cellular energy metabolism and signal transduction, and changes in their composition and content are closely related to the physiological state of cells as well as the occurrence and development of diseases.

With the development of metabolomics technology, untargeted metabolomic analysis of exosomes has become a research hotspot. Untargeted metabolomics can comprehensively detect various metabolites in exosomes without prior knowledge of target metabolites, which can maximize the discovery of potential biomarkers and changes in metabolic pathways. This provides a new perspective for understanding the pathogenesis of diseases, developing new diagnostic methods and therapeutic targets.

Untargeted metabolomic profiling of extracellular vesicles' molecular cargo. Figure 1. Untargeted metabolomics analysis of EVs molecular cargo. (Dudzik D, et al., 2021)

Our Untargeted Exosome Metabolomics Service

Creative Biostructure provides untargeted metabolomics services for exosomes using advanced liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS) technology platforms.

Platform Technology	Features
LC-MS Technology	High sensitivity, high resolution and a wide range of metabolite coverage, which can effectively detect polar and thermally unstable metabolites.
GC-MS Technology	Highly effective for analyzing volatile and thermally stable metabolites.

Our efficient platform can achieve comprehensive capture and accurate quantification of various metabolites in exosomes, including lipids, amino acids, organic acids, vitamins, carbohydrates and their derivatives, as well as nucleosides and nucleotides, ensuring the accuracy and reliability of the detection results.

Untargeted Exosome Metabolomics Analysis Workflow

We provide one-stop exosome untargeted metabolomics analysis services. From sample reception and evaluation to the generation of the final report, we aim to deliver reliable analysis results to customers through strict quality control. The detailed workflow of this experiment is as follows:

Our standard workflow includes the following key steps:

1

Exosome Isolation and Purification

For different types of samples, appropriate methods such as differential centrifugation, density gradient centrifugation, or kit-based methods are used to isolate and purify exosomes, ensuring the purity and integrity of exosomes.

2

Metabolite Extraction

Optimized extraction methods are employed to extract metabolites from the purified exosomes, preserving the structure and activity of various metabolites as much as possible.

3

Mass Spectrometry Detection

The extracted metabolite samples are subjected to LC-MS and GC-MS detection respectively to obtain mass spectrometry data of metabolites.

4

Data Processing and Analysis

Professional bioinformatics software is used for preprocessing the mass spectrometry data, including peak identification, peak matching, quantification, etc. Then multi-omics data analysis is performed, such as screening of differential metabolites, metabolic pathway enrichment analysis, etc.

5

Report Generation

Based on the data analysis results, a detailed service report is generated, which includes experimental methods, metabolite lists, differential analysis results, pathway analysis, etc., and professional interpretation of the results is provided.

Comprehensive untargeted profiling of exosome metabolomics. Figure 2. Comprehensive workflow for untargeted metabolomic profiling of exosomes. (Creative Biostructure)

Supported Sample Range

We support a wide range of samples, including exosomes isolated from cell culture supernatants (e.g., from cancer cells, normal somatic cells), and exosomes from various body fluids such as blood (serum/plasma), urine, saliva, cerebrospinal fluid, and milk.

Sample Submission Guide

Quantity: For cell culture supernatants, provide at least 10 mL. For body fluids, the required volume varies: 200 μL for serum/plasma, 500 μL for urine, 250 μL for saliva, and 200 μL for cerebrospinal fluid.
Storage and Shipping: Store samples at - 80°C immediately after collection. Ship samples on dry ice to ensure the integrity of exosomes and their metabolites during transportation.

Final Delivery

You will receive a comprehensive report that includes:

Identification of metabolites detected in the exosomes.
Quantitative data of metabolite levels.
Statistical analysis results, such as differential metabolite analysis between experimental groups.
Metabolic pathway analysis related to the identified differential metabolites.

Technical Advantages

Exceptional performance: High sensitivity, high resolution, and a broad dynamic range.
Powerful identification capability: Efficient extraction and identification methods enable precise qualitative analysis of a vast array of compounds.
Reliable quality control: A stringent quality control system ensures accurate and dependable qualitative results.
In-depth data analysis: Diversified and systematic bioinformatic analysis solutions are provided.

Case Study

Case: Untargeted Metabolomic Analysis of Serum Exosomes Identifies Potential Biomarkers for Hepatocellular Carcinoma

Background

Exosomes are intimately associated with tumor development and their contents have the potential to serve as highly sensitive tumor-specific markers. This study aims to identify potential biomarkers for hepatocellular carcinoma through comprehensive untargeted metabolomics analysis of exosome samples extracted from the serum of subjects.

Methods

Exosome samples were extracted from the serum of 48 subjects using a commercial kit.
An ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) strategy was used to identify the metabolic compounds.
A total of 18 differential metabolites were identified using the non-targeted metabolomics approach of UPLC-QTOF-MS/MS.

Conclusion

Pathway analysis revealed significant alterations in the arachidonic acid metabolism, linoleic acid metabolism, and unsaturated fatty acid metabolism pathways.
ROC analysis indicated that three metabolites with AUC values exceeding 0.900 were selected as potential biomarkers: caprylic acid and linoleic acid were upregulated in the HCC group, whereas pentadecanoic acid was downregulated.
Linoleic acid, caprylic acid, and pentadecanoic acid are potential biomarkers for diagnosing HCC.

Orthogonal projections to latent structures discriminant analysis score plot. Figure 3. OPLS−DA score plot. (Martín-Blázquez, et al., 2020)

Ready to map the full metabolic fingerprint of your exosomes? Our untargeted LC-MS/GC-MS workflow captures diverse small molecules with rigorous QC and links differential metabolites to pathways and phenotypes through in-depth bioinformatics. At Creative Biostructure, we adapt each module to your sample type and study question and deliver transparent, publication-ready reports you can trust. Contact us to refine your design and receive a fast, tailored quotation.

References

Dudzik D, Macioszek S, Struck-Lewicka W, et al. Perspectives and challenges in extracellular vesicles untargeted metabolomics analysis. TrAC Trends in Analytical Chemistry. 2021, 143: 116382.
Martín-Blázquez, Ariadna, et al. Discovery of pancreatic adenocarcinoma biomarkers by untargeted metabolomics. Cancers. 12.4 (2020): 1002.

Frequently Asked Questions

For any inquiries, our support team is ready to help you get technical support for your research and maximize your experience with Creative Biostructure.

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Lipidomics and Metabolomics in Exosome Research

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