Small Extracellular Vesicles/Exosome Supernatant Preparation

Extracellular Vesicles (EVs) are nanoscale particles secreted by cells, playing a pivotal role in mediating intercellular communication over both short and long distances, and participating in the regulation of various physiological and pathological processes. According to the 2018 guidelines of the International Society for Extracellular Vesicles, EVs are typically classified based on their size. Vesicles with a diameter of 200nm or less are termed small extracellular vesicles (sEVs), often known as exosomes, which are the primary focus of contemporary research. Vesicles larger than 200nm in diameter are categorized as medium or large extracellular vesicles.

 

Plant cells, bacterial cells, and mammalian cells can all produce and secrete extracellular vesicles, which contain proteins, nucleic acids, lipids, and small molecule metabolites. The source of exosomes is mainly separated from the supernatant collected through cell culture, tissue enzymatic hydrolysis, body fluid acquisition, etc. NextGen Exsome possesses extensive experience in sample processing. In the upstream cell culture stage, we can provide customized processing plans. We also select different enzymatic digestion or juice extraction methods for tissues of varying density. Additionally, we can perform special pretreatments for viscous or solid-liquid mixed samples to facilitate the subsequent separation of exosomes.

 

Cultivation

During the isolation and purification of exosomes, cell culture supernatant is a common sample source. The origin, culture duration, and conditions of the sample significantly influence the quantity and quality of the exosomes. Thus, standardizing and uniformly processing the sample source prior to experimentation is essential. Furthermore, the composition of exosomes secreted by different cells may vary, it is necessary to select the appropriate cell source based on the specific objectives of the experiment.

 

Microcarrier Cultivation

NextGen Exsome using microcarriers that composed of tens of thousands of elastic, three-dimensional porous microcarriers, with a porosity rate of over 90%. The particle size can be precisely controlled between 50-500 micrometers, with a uniformity superior to 100 micrometers, thereby creating a true 3D biomimetic culture environment. These microcarriers utilize a specialized cleavage technology, enabling a 100% cell harvest rate. Residues from the microcarriers and lysis have obtained the residue detection report and related safety evaluation report from the National Institutes for Food and Drug Control. This product has obtained two CDE excipient qualifications and one FDA-DMF excipient qualification, making it suitable for the scaled-up culture of most adherent cells and the harvest of cells, viruses, and cell products.

 

Enzymatic Digestion

Enzymatic digestion is an efficient method for the breakdown of biological tissues, widely used in the extraction and separation of exosomes. This technique effectively reduces tissue samples to single-cell suspensions, which after the removal of cellular debris and tissue fragments, allows the extraction of high-purity exosomes from the supernatant. Animal tissues such as brain, adipose, skin, liver, and bone tissues can be digested using a combination of various enzymes (like trypsin, protease, collagenase, and dissociation enzymes, ect.) to achieve optimal supernatant preparation. Similarly, plant tissues (such as roots, stems, leaves, flowers, fruits, and seeds) can be digested with plant-specific enzymes (like pectinase, cellulase, etc.).

 

Services at NextGen Exsome

1.Cell Culture by Culture Dishes

Cell culture is performed utilizing the standard culture dish technique with either serum-free or exosome-depleted serum culture medium. This method is optimal for small-scale collection (50-200ml).

 

Deliverables:Cell supernatant.

 

2.Cell Culture by Microcarrier

Cell culture is executed using the microcarrier suspension approach, employing serum-free or exosome-depleted serum culture mediums, making it ideal for large-scale collection (200-5000ml).

 

Deliverables:Cell supernatant.

 

3.Animal Tissue Enzymatic Hydrolysis

Selecting an appropriate enzymatic solution system (such as trypsin, protease, collagenase, dispase, etc.) for tissue digestion.

 

Deliverables:Digestion solution supernatant.

 

4.Plant Tissue Enzymatic Hydrolysis

Selecting an appropriate enzymatic solution system (e.g., pectinase, cellulase, etc.) for tissue digestion.

 

Deliverables:Digestion solution supernatant.