ExoPureTM Immunobeads (Overall Exosome Isolation, cell media)
Usage: RUO Cat: M1030-10, -20 assays; Store at 4°C
Introduction
Exosomes are small endosome derived lipid nanoparticles (50-120 nm) actively secreted by exocytosis by most living cells
Exosome release occurs either constitutively or upon induction, under both normal and pathological conditions, in a dynamic, regulated and functionally relevant manner. Both the amount and molecular composition of the released exosomes depend on the state of a parent cell.
Exosomes have been isolated from diverse cell lines (hematopoietic cells, tumor lines, primary cultures, and virus infected cells) as well as from biological fluids in particular blood (e.g. serum and plasma from cancer patients) and other body fluids (broncho alveolar lavage fluid, pleural effusions, synovial fluid, urine, amniotic fluid, semen, saliva etc). Exosomes have pleiotropic physiological and pathological functions and an emerging role in diverse pathological conditions such as cancer, infectious and neurodegenerative diseases.
Biovision offers ExoPureTM Immunobeads are for capturing and isolating overall or specific exosome sub-populations. Latex immunobeads are covalently coupled with antibodies against exosome surface antigens, allowing exosome capture from human biofluids (tested for plasma, serum and urine) and cell culture media without pre-purification steps (ultracentrifuge or other methods for exosome purification).
Biovision immunobeads are able to capture the overall exosome population (Immunobeads for Overall Exosome capture) or to enrich exosome subpopulation derived from a tumoral source (Tumor-derived exosome capture and enrichment).
Immunobeads are supplied with an Exosome Elution Buffer, that allows detachment and elution of captured exosomes for downstream analyses, and with a Bead Regeneration Buffer to regenerate immunobeads for further usage.
All latex Immunobeads are available in two sizes (0.4 and 1 micron of diameter) and are sold in packages of 10 and 20 reactions. The latex immunobeads are pre-coated with anti-mouse or anti-Rabbit antibodies. It uses an easy, fast and efficient protocol.
Beads are mixed together with the sample and incubated overnight (ON) for exosomes binding. After ON incubation beads can be easily recovered by centrifugation and washed with appropriate buffer.
Bead pellet can be directly used for nucleic acid extraction or protein analysis. Alternatively, exosomes can be eluted off the beads with theExosomeElutionBuffer and used forfunctional studies.
Application
- For overall exosome isolation from cell culture media.
- Small sample volume of cell culture medium.
- No ultracentrifugation or other methods for exosome purification required.
- Immunobeads are ready to use and stable for long term (up to 8 months).
- Immunobeads are suitable for nucleic acid extraction, protein profiling of exosome markers, exosome elution from immunobeads.
- A mouse monoclonal antibody has been used for coating.
Sample Type
- Cell culture media.
- Concentrated (10X) cell culturemedium samples is recommended prior capture according to our suggested protocol.
Kit Contents
| Components | M1030-10 | M1030-20 |
|---|---|---|
| Pre-coupled Latex Immunobeads | 100 µl (10 reactions) | 200 µl (20 reactions) |
| Exosome Elution Buffer | 1 bottle (650 µl) | 1 bottle (450 µl) |
| Bead Regeneration Buffer | 1 bottle (10 ml) | 1 bottle (10 ml) |
ExoPureTM Immunobeads Assay Protocol
- Human Cell culture medium sample preparation
- 10 min at 300g
- 20 min at 1600g
- 30 min at 10,000g. Concentrate cell supernatant 10X (from 10 ml to 1 ml) in spin concentrator
- The quantity of exosomes could vary between samples. Concentration factors are given for information purposes only, a larger starting amount of sample should be used if the signal is weak.
- Exosome immunocapture: Add
10 µl of pre-coupled beads to 0.5 ml up to 1 mlof biological sample (plasma, urine or cell culture supernatant previously precleared). Incubate overnight at 4°C in rotator. Remark: Incubation can be carried out also at room temperature for at least 4 hr in rotator. After exosome binding wash beads twice with 1 ml of PBS resuspending up and down. 10- 15 times. In each step remove the supernatant by centrifugation at 5000g for 10 min. The prepared beads can be used for further captured exosome characterization including both protein and nucleic acid content analysis, or exosomes can be recovered and analyzed.