RNA sample should be submitted in RNase-free water, RNA Stabilization Reagent, or 10mM Tris pH 8.0. All total RNA samples should be DNA-free. RNA samples require an OD A260/A280 ratio ≥ 1.8, A260/230 ratio≥ 1.8 and a RIN ≥ 6. Ship with dry ice.
*Due to varying requirements for different samples, please consult our technical team for additional guidelines based on your projects.
*If you want to explore and inquire further about our RNA sequencing sample requirements and preprocessing procedures, kindly refer to our Service Sample Submission Guidelines for comprehensive information.
EXOSOME PREPARATION GUIDELINES
Blood
For optimal results, serum or plasma should be separated promptly after blood collection. Whenever possible, choose plasma to minimize the exosomal effects of platelet origin.
Plasma
Use a blood collection needle and an EDTA anticoagulant tube to draw whole blood. Gently mix and store at room temperature or 4°C, proceeding to the next step within 1 hour.
Centrifuge the blood at 4°C for 10 minutes at 1900 g. Carefully aspirate the supernatant as plasma, discarding the last 500 μl or so. Centrifuge the obtained plasma again at 4°C, 3000 g for 15 minutes, carefully aspirating without touching the sediment.
Freeze the plasma at -80°C.
Tip: Send a sample volume of 4 ml or more (approximately 8-10 ml of whole blood is needed to separate 4 ml of plasma).
Serum
Collect 10 ml of blood with a blood collection needle and an ordinary serum tube (without any reagents, 10 ml size).
Allow it to stand at room temperature for 30 minutes, then at 4°C for 3-4 hours, allowing visible blood clots to precipitate.
Use a pipette to collect the yellowish serum (approximately 4 ml) into a 15 ml centrifuge tube. Centrifuge at 3000 g for 15 minutes at 4°C, carefully transferring the supernatant to a new tube for optimal serum quality.
Freeze the centrifuged serum in the refrigerator at -80°C within 15 minutes.
Tip: Send more than 4 ml of the sample (approximately 10-15 ml of whole blood is needed to separate 4 ml of serum).
Cell Supernatant
Cell Culture Adherent cells: Culture in normal serum-containing medium until reaching 70%-80% confluence. Suspension cells: Achieve a cell density of 60%-70%.
Medium Replacement or Cell Collection Adherent cells: Replace the original medium with exosome-free or serum-free medium. Suspension cells: Collect cells at 300 g, 4°C, for 10 minutes.
Continued Culture Suspend cells in exosome-free or serum-free medium and continue culturing for 24-48 hours, determining the collection time based on cell growth rate.
Initial Centrifugation Centrifuge at 300 g, 4°C, for 10 minutes. Carefully aspirate the supernatant, avoiding cell or debris contamination.
Secondary Centrifugation Centrifuge the collected supernatant at 3000 g, 4°C, for 15 minutes. Ensure thorough removal of cells or cellular debris.
Sample Handling Collect the clarified supernatant with caution to prevent cell or debris contamination. Combine identical cell culture supernatant samples and transfer them into sterile containers. Short-term storage (1-2 days) at 4°C is acceptable. For long-term storage, freeze at -80°C. Note that storage at both temperatures may impact yield, and it's recommended to proceed with exosome isolation promptly after separation. Suggested Volume: ≥ 20 ml.
Urine
Collection Collect anterior/middle and posterior morning urine, aiming for approximately 100 ml of "fresh" urine from the subject. Be mindful to avoid bacterial contamination. Control diet before collection. Temporarily store at 4°C (for no more than 8 hours).
Centrifugation Centrifuge the urine at 3000×g for 15 minutes to effectively remove cells or cell debris.
Storage Store the clarified urine in the refrigerator at -80°C. Suggested Sample Volume: ≥ 20 ml.
Cerebrospinal Fluid (CSF)
Collection Collect 10 ml of cerebrospinal fluid via lumbar puncture. Once separated, promptly place the sample on ice or at 4°C for a brief period (not exceeding 4 hours). To prevent blood contamination, refrain from using sampling tubes containing heparin anticoagulant.
Centrifugation Centrifuge the collected cerebrospinal fluid at 3000×g for 15 minutes to eliminate cells or cell debris.
Storage Store the clarified cerebrospinal fluid in the refrigerator at -80°C. Suggested Sample Size: ≥ 10 ml.
Chest Fluid
Collection and Temporary Storage After clinical collection of pleural fluid, if immediate processing is not possible, temporarily store it at 4°C (for not more than 8 hours).
Initial Centrifugation Centrifuge the collected pleural fluid at 1000 g for 10 minutes. Collect the resulting supernatant.
Secondary Centrifugation Centrifuge the supernatant from step 2 at 3000 g for 10 minutes. Collect the resulting supernatant.
Storage Store the clarified pleural fluid in the refrigerator at -80°C. Suggested Sample Volume: ≥ 30 ml.
Ascites Sample
Collection and Temporary Storage If ascites cannot be processed immediately after clinical collection, store it temporarily at 4°C (for not more than 8 hours).
Centrifugation for Residue Removal Centrifuge the collected ascites at 2000 g for 20 minutes at 4°C (room temperature is also acceptable) to effectively remove residue in the ascites.
Storage Store the processed ascites in the refrigerator at -80°C. Suggested Sample Size: ≥ 30 ml.
Bile Sample
Collection Collect bile in a sterile container.
Centrifugation for Sediment Removal Centrifuge the collected bile at 3000 g for 10 minutes at 4°C to eliminate cellular sediment and debris.
Supernatant Collection and Storage: Collect the supernatant from the centrifuged bile and store it at 4°C or -20°C for long-term preservation. Suggested Sample Size: ≥ 5 ml.
* For Research Use Only. Not for use in diagnostic procedures.
