2 best Reasons to use our kits to purify exosome
95% exosome purity isolation
10 fold higher exosome yield than all other kits
Our Exosome kits help avoid 80% Exosome waste in your sample.
Product Application & Flow Chart: (click on products to show/hide description)
P100 PureExo Cell: cell media exosome
Product Size: 10 reactions (2 mL~4 mL medium / reaction)
Yield: typically 50~200 µL exosome can be isolated from each reaction. If use our P200 kit in the next step, 150 ~ 400 µg exosomal protein or 50 ~ 200 ng exosomal RNA can be extracted from 50~200 µL exosome
Storage: keep all bottles upright, in cool and dark place.
Product Description
Our PureExo® Exosome Isolation kit (for cell culture media) enable fast and efficient recovery of intact exosomes from cell culture media.
- No ultra-centrifugation and tedious isolation procedures (< 2 hours);
- Better recovery of intact exosomes than ultra-centrifugation: 5-10 folds yield;
- Save cost than antibodies-beads method;
- Isolate Pure exosome;
- Use as little as 2 mL cell media to achieve high yield of exosomes for any downstream applications: EM study, exosome label, exosome subpopulation, qRT-PCR profiling of exosomal miRNAs, and gel analysis of exosomal proteins.
Product Contents (of full unit package)
Component | Amount (full unit) | Storage |
---|---|---|
Solution A (blue) | 2.5 mL | Room temperature |
Solution B | 2.5 mL | Room temperature |
Solution C * | 10 mL | Room temperature |
PureExo® Columns | 10 x (1.5 mL) | Room temperature |
Reaction Volume Table (important):
Cell culture media | Mixture A/B/C | = | Solution A | + | Solution B | + | Solution C |
---|---|---|---|---|---|---|---|
2 mL (min / Rxn) | 0.75 mL | = | 0.125 mL | + | 0.125 mL | + | 0.5 mL |
3 mL | 1.125 mL | = | 0.187 mL | + | 0.187 mL | + | 0.75 mL |
4 mL (max / Rxn) | 1.5 mL | = | 0.25 mL | + | 0.25 mL | + | 1 mL |
- The maximum medium volume of each reaction is 4 mL from at most 5 x 105 cells. One PureExo® Columns per reaction.Do not exceed the suggested sample volume or the cell number. Otherwise it will cause indistinct layer separation and column clog.
- If the cultured cells are highly proliferative cell, such as tumor cells or stem cells, reduce loading amount of cell culture medium by half to process.
Protocol (example of processing 2 mL medium)
- 1.Collect 2 mL cell culture medium. If the culture medium is from Bioreactor system, dilute the medium to less than
2 x 105 cells / 2 mL. - 2.Centrifuge the cell media at 600× g for 10 minutes at 4°C to remove cells or debris.
- Important: skip this step may cause filter clog in step 14.
- 3.Transfer clear supernatant (cell-free culture media) to a new glass tube 1 and keep it on ice.
- 4.In glass tube 2, add the solutions in the following order to prepare mixture A/B/C:
- 5.Vortex the tube 2 (mixture A/B/C) for 5-10 seconds to obtain a homogenous solution.
- 6.Add tube 2 (mixture A/B/C 0.75 mL) to tube 1 (2 mL cell-free culture media).
- 7.Tightly cap tube 1, vigorously vortex for 30 seconds, then incubate at 4°C for 1 hours.
- 8a.The mixture now appear as 3 layers (as shown in the following figure):
- 8b.Sometimes, only two layers (medium color layer and white cloudy / fluffy layer) are visible (as shown in the following figure):
- 9.Transfer the left over in the tube to a new Eppendorf tube and spin at 1,000x g for 3 minutes. A new three-layer separation will occur (top medium color layer, middle fluff layer and bottom colorless layer as shown in the following figure).
- 10.Pipet out and discard the top layer. Insert pipette tip down to the tube bottom to remove the colorless bottom layercompletely. Therefore only the “fluff” is left in the tube.
- 11.Repeat step 9 and step 10 once.
- 12.Leave the Eppendorf tube cap open to air dry for 5-10 minutes at room temp. (do not over dry).
- 13.Add 1× PBS as much as 1-2 volumes of the collected fluff to the tube, and resuspend the “fluff” by pipetting up and downvigorously.
- 14.Transfer the suspension carefully into PureExo® Column (provided) and spin the Column at 2,000× g for 5 minutes to collect all the flow-through;
- 15.The “flow-through” is the isolated pure exosome (exosome suspended in PBS). Use the isolated pure exosome directly or store at 4°C for up to 1 week, or at ≤80°C for up to 3 months.
1stSolution A (blue) | 0.125 mL |
2ndSolution B | 0.125 mL |
3rdSolution C * | 0.5 mL |
Carefully aspirate the top layer using a pipette without disturbing the middle fluff layer and discard it, then go to step 9.
Remove and discard the top layer, then go to step 9.
More info: Exosome Isolation kit (for cell culture media)
Manual (protocol): Download
Price: 310 Euro
P101 PureExo Serum: serum exosome
Product Size: 10 reactions (100~600 µL serum / reaction)
Yield: if start with 200 µL serum, typically 100~200 µL exosome can be isolated. If use our P200 kit in the next step, 300 ~ 400 µg exosomal protein or 200 ~ 300 ng exosomal RNA can be extracted.
Storage: keep all bottles upright, in cool and dark place.
Product Description
Our PureExo® Exosome Isolation kit (for serum) enable fast and efficient recovery of intact exosomes from serum.
- No ultra-centrifugation and tedious isolation procedures (<2 hours);
- Better recovery of intact exosomes than ultra-centrifugation: 5-10 folds yield;
- Save cost than antibodies-beads method;
- Isolate Pure exosomes;
- Use as little as 100 µl serum to achieve high yield of exosomes for any downstream applications: including EM study, exosome label, exosome subpopulation, qRT-PCR profiling of exosomal miRNAs, and gel analysis of exosomal proteins.
Product Contents (of full unit package)
This kit contains reagents sufficient for processing 6 mL of serum.
Component | Amount (full unit) | Storage |
---|---|---|
Solution A (orange) | 1 mL | Room temperature |
Solution B | 1 mL | Room temperature |
Solution C * | 4 mL | Room temperature |
PureExo® Columns | 10 x (1.5 mL) | Room temperature |
Reaction volumes table (Important)
Serum | Mixture A/B/C | = | Solution A | + | Solution B | + | Solution C |
---|---|---|---|---|---|---|---|
100 µl | 100 µl | = | 17 µl | + | 17 µl | + | 66 µl |
200 µl | 200 µl | = | 33 µl | + | 33 µl | + | 134 µl |
300 µl | 300 µl | = | 50 µl | + | 50 µl | + | 200 µl |
400 µl | 400 µl | = | 67 µl | + | 67 µl | + | 266 µl |
500 µl | 500 µl | = | 83 µl | + | 83 µl | + | 334 µl |
600 µl | 600 µl | = | 100 µl | + | 100 µl | + | 400 µl |
- The maximum serum volume of reaction is 600 µl. One PureExo® Columns per reaction. Do not exceed the suggested sample volume, otherwise it will cause indistinct layer separation and column clog.
Protocol (example for processing 100 µl serum)
- 1.Collect the serum sample and keep it on ice. Or thaw the frozen sample completely at room temperature, and keep it on ice.
- 2.Centrifuge the serum sample at 2,000× g for 10 minutes at 4℃ to remove debris.
- Important: skip this step may cause filter clog in step 14.
- 3.Transfer the clear supernatant to glass tube 1 without disturbing the pellet, and keep it on ice.
- 4.In glass tube 2, add the solutions in the following order to prepare mixture A/B/C:
- 5.Vortex the tube 2 (mixture A/B/C) for 5-10 seconds to obtain a homogenous solution.
- 6.Add tube 2 (100 µl mixture A/B/C) to tube 1 (100 µl serum).
- 7.Tightly cap tube 1, vigorously vortex for 30 seconds, then incubate at 4°C for 1 hours.
- 8a.The mixture now appear as 3 layers:
- 8b.Sometimes, only two layers (orange color top layer and cloudy bottom layer) are visible, remove and discard the top layer. Go to step 9.
- 9.Transfer the left over in the tube to a new Eppendorf tube and spin at 1,000x g for 3 minutes. A new three-layer separation will occur: top orange color layer, middle fluff layer and bottom colorless layer. (As shown in the following figure.)
- 10.Pipet out and discard the top layer. Insert pipette tip down to the tube bottom to remove the colorless bottom layer completely. Therefore only the “fluff” is left in the tube.
- 11.Repeat step 9 and step 10 once.
- 12.Leave the Eppendorf tube cap open to air dry for 5-10 minutes at room temp. (do not over dry).
- 13.Add 1× PBS as much as 1-2 volumes of the collected fluff to the tube, and resuspend the “fluff” by pipetting up and downvigorously.
- 14.Transfer the suspension carefully into PureExo® Column (provided) and spin the Column at 2,000× g for 5 minutes to collect all the flow-through.
- 15.The “flow-through” is the isolated pure exosome (exosome suspended in PBS). Use the isolated pure exosome directly or store at 4°C for up to 1 week, or at ≤80°C for up to 3 months.
1stSolution A (orange) | 17 µl |
2ndSolution B | 17 µl |
3rdSolution C * | 66 µl |
Aspirate the top layer using a pipette without disturbing the middle fluff layer and discard it. Go to step 9.
More info: Exosome Isolation kit (for serum)
Manual (protocol): Download
Price: 310 Euro
P120 DiagExo Urine: urine exosome
Product Size: 20 reactions (3 mL urine / reaction)
Yield: typically 50~200 µL exosome can be isolated from each reaction. If use our P200 kit in the next step, 150 ~ 400 µg exosomal protein or 50 ~ 200 ng exosomal RNA can be extracted from 50~200 µL exosome.
Storage: keep all bottles upright, in cool and dark place.
Application: to isolate exosome from urine.
Product Description
Exosomes are cell-derived extracellular vesicles with a diameter of 20-200 nm and present in a wide range of biological fluids including urine, blood, amniotic fluid, saliva, peritoneal fluid, inflammatory fluid, and breast milk. Exosomes play important roles in intercellular communications and can potentially be used for prognosis, therapy, and biomarkers for some human diseases.
Our DiagExo® Urinary Exosome Isolation kit enable fast and efficient recovery of intact exosomes from urine.
- Isolate Pure exosome;
- 10 folds yield: Better recovery of intact exosomes than ultra-centrifugation
- No ultra-centrifugation and tedious isolation procedures (< 2 hours);
- Save cost vs. antibodies-beads method;
- Use as little as 1 mL urine to achieve high yield of exosomes for any downstream applications: EM study, exosome labeling, exosome subpopulation, qRT-PCR profiling of exosomal miRNA, ELISA and gel analysis of exosomal proteins.
Product Contents
Component | Amount | Storage |
---|---|---|
Solution A (blue) | 5 mL | room temperature |
Solution B | 5 mL | room temperature |
Solution C * | 20 mL | room temperature |
DiagExo® Exosome Columns | 20 x (1.5 mL) | room temperature |
Reaction Volume Table (important):
Volume of urine | Mixture A/B/C | = | Solution A | + | Solution B | + | Solution C |
---|---|---|---|---|---|---|---|
1 mL (min / Rxn) | 0.5 mL | = | 0.083 mL | + | 0.083 mL | + | 0.334 mL |
2 mL | 1.0 mL | = | 0.166 mL | + | 0.166 mL | + | 0.668 mL |
3 mL (max / Rxn) | 1.5 mL | = | 0.25 mL | + | 0.25 mL | + | 1 mL |
The suggested maximum urine volume of each reaction is 3 mL. One PureExo® Columns per reaction. Loading amount of urine exceeds the suggested sample volume will cause indistinct layer separation and column clog.
Protocol (example for processing 2 mL urine)
- Sample prepare: Centrifuge the 2 mL urine sample at 2,000× g for 10 minutes at room temperature to remove cells and debris;
- Without disturbing pellets, transfer the clear supernatant to a new glass tube 1 and keep it on ice
- In glass tube 2, add the solutions in the following order to prepare mixture A/B/C:
- Vortex the tube 2 (mixture A/B/C) for 5-10 seconds to obtain a homogenous solution
- Add tube 2 (mixture A/B/C 1.0 mL) to tube 1 (2 mL urine).
- Tightly cap tube 1, vigorously vortex for 30 seconds, then incubate at 4°C for 1 hour
-
7a.The mixture now appear as 3 layers (as shown in figure bollow):
Carefully, aspirate the top layer using a pipette without disturbing the middle fluff layer and discard it. Then go to step 9.
7b.Sometimes, only two layers (blue color top layer and white fluffy bottom layer) are visible (as shown in the following figure). Carefully remove / aspirate the top layer and discard it. Then go to step 9.
-
Transfer the left over in the tube to a new Eppendorf tube and spin at 1,000x g for 3 minutes. A new three-layer separation will occur (top blue color layer, middle fluff layer and bottom colorless layer as shown in the following figure).
-
Pipet out and discard the top layer. Insert pipette tip down to the tube bottom to remove the colorless bottom layer completely. Therefore only the “fluff” is left in the tube.
- Repeat step 9 and step 10 once.
- Leave the Eppendorf tube cap open to air dry for 5-10 min. at room temp (do not over dry).
- Add 1× PBS as much as 1-2 volumes of the collected fluff to re-suspend the “fluff” by pipetting up and down vigorously.
- Transfer the suspension carefully into PureExo® Column (provided) and spin the Column at 2,000× g for 5 minutes to collect all the flow-through.
- The “flow-through” is the isolated pure exosome (exosome suspended in PBS). Use the isolated pure exosome directly or store at 4°C for up to 1 week, or at ≤ 80°C for up to 3 months.
1stSolution A (blue) | 0.166 mL |
2ndSolution B | 0.166 mL |
3rdSolution C * | 0.668 mL |
More info: Urinary Exosome Isolation kit
Manual (protocol): Download
Price: 566 Euro
P121 DiagExo BodyFluid: body fluid exosome
Product Size: 20 reactions (0.1 ~ 2 mL body fluid / reaction). The yield of exosome varies depending on the different sample type.
Storage: keep all bottles upright, in cool and dark place.
Application: For isolating exosome from, Cerebrospinal fluid (CSF), Amniotic fluid, Inflammatory fluid, Lymph fluid, Breast milk, Saliva, Gastrointestinal fluid (GI), and Broncho alveolar lavage fluid.
Product Description
Use 0.5 - 2 mL human body fluid to achieve high yield of exosomes for any downstream applications: EM study, exosome labeling, exosome subpopulation, qRT-PCR profiling of exosomal miRNA, ELISA and gel analysis of exosomal proteins.
Product Contents
Component | Amount | Storage |
---|---|---|
Solution A (orange) | 2 mL | room temperature |
Solution B | 2 mL | room temperature |
Solution C * | 8 mL | room temperature |
DiagExo® Exosome Columns | 20 x (1.5 mL) | room temperature |
Reaction Volume Table (important):
Solution A/B/C | Serum | CSF | Amniotic fluid | Inflamma-tory fluid | Lymph fluid | Breast milk | Saliva | GI fluid | Broncho alveolar lavage fluid |
---|---|---|---|---|---|---|---|---|---|
Solution A20 ul Solution B20 ul Solution C80 ul |
100 ul | 100 ul | 100 ul | 100 ul | 100 ul | 200 ul | 400 ul | 400 ul | 400 ul |
Solution A40 ul Solution B40 ul Solution C160 ul |
200 ul | 200 ul | 200 ul | 200 ul | 200 ul | 400 ul | 800 ul | 800 ul | 800 ul |
Solution A60 ul Solution B60 ul Solution C240 ul |
300 ul | 300 ul | 300 ul | 300 ul | 300 ul | 600 ul | 1.2 mL | 1.2 mL | 1.2 mL |
Solution A80 ul Solution B80 ul Solution C320 ul |
400 ul | 400 ul | 400 ul | 400 ul | 400 ul | 800 ul | 1.6 mL | 1.6 mL | 1.6 mL |
Solution A100 ul Solution B100 ul Solution C400 ul |
500 ul | 500 ul | 500 ul | 500 ul | 500 ul | 1 mL | 2 mL | 2 mL | 2 mL |
Protocol (example for processing 100 ul of serum)
- Sample prepare: Collect suggested volume of specific body fluid indicated in the above table. If start with frozen sample(s), thaw the frozen sample completely at room temperature, and keep it on ice.
- Centrifuge at 2,000× g for 10 minutes at room temperature to remove cells and debris.
- Without disturbing pellets, transfer clear supernatant to a new glass tube 1 and leave it on ice till isolation;
- Prepare mixture of A/B/C: In glass tube 2, add solutions in the following order:
- Vortex gently tube 2 (mixture A/B/C) for 5 - 10 seconds to obtain a homogenous solution;
- Add tube 2 (mixture A/B/C) to tube 1 (body fluid sample);
- Tightly cap tube 1, vigorously vortex for 30 seconds, then incubate at 4°C for 1 hour.
-
8a.
The mixture now appear as 3 layers (as shown in the following figure):
Carefully, aspirate the top layer using a pipette without disturbing the middle fluff layer and discard it. Go to step 9.
8b.Sometimes, only two layers (orange color top layer and cloudy bottom layer) are visible, remove and discard the top layer. Go to step 9.
-
Transfer the left over in the tube to a new Eppendorf tube and spin at 1,000x g for 3 minutes. A new three-layer separation will occur: top orange color layer, middle fluff layer and bottom colorless layer (as shown in the following figure).
-
Pipet out and discard the top layer. Insert pipette tip down to the tube bottom to remove the colorless bottom layer completely. Therefore only the “fluff” is left in the tube.
- Repeat step 9 and step 10 once.
- Leave the Eppendorf tube cap open to air dry for 5-10 minutes at room temp. (Do not over dry).
- Add 1× PBS as much as 1-2 volumes of the collected fluff to the tube, and resuspsend the “fluff” by pipetting up and downvigorously.
- Transfer the suspension carefully into PureExo® Column (provided) and spin the Column at 2,000× g for 5 minutes to collect all the flow-through.
- The “flow-through” is the isolated pure exosome (exosome suspended in PBS). Use the isolated pure exosome directly or store at 4°C for up to 1 week, or at ≤80°C for up to 3 months.
1stSolution A (orange) | 20 ul |
2ndSolution B | 20 ul |
3rdSolution C * | 80 ul |
Remark:
For human body fluid sample volume other than the suggested volume in the table, scale up or down the solution A, B and Cproportionally.
The PH value in the body fluids will not affect our exosome isolation efficiency.
More info: Human Body Fluid Exosome Isolation kit
Manual (protocol): Download
Price: 566 Euro
P200 RNA/Protein Kit: exosome RNA / Protein
Storage: keep all bottles upright, in cool and dark place.
Application: our “2 in 1” kit is for extracting both exosomal RNA and exosomal protein from pure exosome isolated by our Exosome Isolation Kits (Cat.#: P100, P101, D100, D101).
Product Size: 20 extractions
Product Contents
Component | Amount | Storage |
---|---|---|
Exosomal Protein Lysis buffer * | 2 mL | -20C |
N1 | 5 mL | room temperature |
N2 | 1 mL | room temperature |
N3 | 2.5 mL | room temperature |
N4 | 10 mL | room temperature |
RNA elution buffer | 0.5 mL | room temperature |
Important: RNA is sensitive to RNase. Before starting RNA extraction, prepare clean lab bench and wipe working surface and pipettors with RNase decontamination solution, such as Ambion® RNaseZap®. Always wear clean laboratory gloves during manipulation.
Protocol
Sample prepare
- Transfer the isolated exosomes (by our PureExo® Exosome Isolation kits) to an RNase free tube. Add 1× PBS buffer to the exosomes to a final total volume of 100 ul.
- Split the exosomes into two portions: 75 µl for RNA extraction and 25 µl for protein extraction, if both RNA and protein extraction are desired.
- Transfer the 75 µl exosomes to an RNase free tube and add 250 μl N1; mix extensively by pipetting up and down and incubate 5 min at room temp.
* For other volumes of exosome, adjust N1 volume proportionally. - Add 50μl N2 to the sample and vortex vigorously for 15 seconds and incubate at room temp for 2-3 min.
- Centrifuge sample at 12,000x g for 15 min. at 4 C.
- Without disturbing interphase, transfer the upper aqueous phase to a new RNase free tube.
- In the new RNase free tube, add 125 μl N3 to precipitate exosomal RNA;
- Incubate for 15 min. at room temperature. Centrifuge at 12,000x g for 10 min. at 4 C;
- The exosomal RNA precipitates as gel like pellet on the bottom / side of the tube. Carefully remove / discard the supernatant.
- Wash RNA pellet with 250 μl N4, mix and centrifuge at 7,500x g for 5 min. at 4 C. Remove supernatant without disturbing RNA pellet;
- Repeat step 10 once;
- Air dry the exosomal RNA pellet for 10 min. at room temp;
- Dissolve the exosomal RNA pellet in 10-15 μl RNA Elution buffer. Use this extracted exosomal RNA for downstream assay or store it at -80 C for future use.
- Thaw exosomal protein lysis buffer aliquot at room temperature and keep it on ice.
- Transfer the 25 µl exosome sample to a clean tube and add exosomal protein lysis buffer 50-100 μl; mix the sample well by pipetting up and down.
- Incubate 15 min. at 4 C and centrifuge the sample at 14,000x g for 10 min. at 4 C.
- The supernatant is the extracted exosomal protein. Transfer the supernatant to a clean tube and keep on ice. Measure the protein concentration. Use it for downstream assay or store the samples at -80 C for up to 3 months.
Exosomal RNA extraction
Homogenization
Phase Separation
Precipitation
Wash
Elution
Exosomal protein extraction
More info: Exosomal RNA and Protein Extraction Kit
Manual (protocol): Download
Price: 195 Euro
P300 Serum ExoProtein: serum Protein
Storage: keep all bottles upright, in cool and dark place.
Application: For directly isolating exosomal protein from human serum, breast milk, saliva, peritoneal fluid, cerebrospinal fluid (CSF), GI fluid, and amniotic fluid
Product Size: 10 reactions
Product Description:
Our DiagExo® Serum Exosomal Protein Extraction kit enables fast and efficient extraction of exosomal proteins from as little as 100 ul serum. The high yield of exosomal proteins for further downstream applications: ELISA, protein mass spectrometry, protein biomarker verification, gel analysis of exosomal proteins, etc. It also works for similar volume of breast milk, saliva, peritoneal fluid, cerebrospinal fluid, lymph fluid, GI fluid, and amniotic fluid.
Product Contents: (10 reactions)
Component | Amount | Storage |
---|---|---|
Solution A (orange) | 1 mL | room temperature |
Solution B | 1 mL | room temperature |
Solution C* | 4 mL | room temperature |
DiagExo® Columns | 10 (1.5 mL) | room temperature |
DiagExo® lysis buffer | 1 mL | 4℃ or -20°C ** |
* Cap the Solution C bottle immediately after each use.
** Short term (up to 7 days) store at 2–8°C. Long term, aliquot and store at -20°C
Reaction Volume Table (important)
Suggested volume | Serum | CSF | Amniotic fluid | Inflamma-tory fluid | Lymph fluid | Breast milk | Saliva | GI fluid | Broncho alveolar lavage fluid |
---|---|---|---|---|---|---|---|---|---|
Solution A20 ul Solution B20 ul Solution C80 ul |
100 ul | 100 ul | 100 ul | 100 ul | 100 ul | 200 ul | 400 ul | 400 ul | 400 ul |
Solution A40 ul Solution B40 ul Solution C160 ul |
200 ul | 200 ul | 200 ul | 200 ul | 200 ul | 400 ul | 800 ul | 800 ul | 800 ul |
Solution A60 ul Solution B60 ul Solution C240 ul |
300 ul | 300 ul | 300 ul | 300 ul | 300 ul | 600 ul | 1.2 mL | 1.2 mL | 1.2 mL |
Solution A80 ul Solution B80 ul Solution C320 ul |
400 ul | 400 ul | 400 ul | 400 ul | 400 ul | 800 ul | 1.6 mL | 1.6 mL | 1.6 mL |
Solution A100 ul Solution B100 ul Solution C400 ul |
500 ul | 500 ul | 500 ul | 500 ul | 500 ul | 1 mL | 2 mL | 2 mL | 2 mL |
Protocol (example of processing 100 ul to 0.5 mL sample of body fluids as listed in the table. Using higher volume than listed in the table may cause indistinct layer separation and column clog.)
- Collect body fluid samples, and keep on ice. If you start with frozen samples, thaw them completely at room temperature, and then keep the samples on ice.
- Centrifuge the sample at 2,000× g for 10 minutes at 4°C to remove cells and debris.
- Without disturbing the pellet, transfer the clear supernatant to a new glass tube 1, and keep it on. Please refer to the “Reaction Volume Table” for the ratio of sample volume vs. solution A/B/C.
- Prepare mixture of A/B/C: In glass tube 2, according to the “Reaction Volume Table”, add solutions in the following order:
- Vortex gently tube 2 (mixture A/B/C) 5 - 10 seconds to obtain a homogenous solution.
- Add tube 2 (mixture A/B/C) to tube 1 (body fluid sample).
- Tightly cap tube 1, vigorously vortex for 30 seconds, then incubate at 4°C for 1 hour.
-
8a.
The mixture now appear as 3 layers:
Carefully, aspirate the top layer using a pipette without disturbing the middle fluff layer and discard it. Go to step 9.8b.Sometimes, only two layers (orange color top layer and cloudy bottom layer) are visible, remove and discard the top layer. Go to step 9.
-
Transfer the left over in the tube to a new Eppendorf tube and spin at 1,000× g for 3 minutes. A new three-layer separation will occur: top orange color layer, middle fluff layer and bottom colorless layer. (See figure on next page for detail.)
-
Pipet out and discard the top layer. Insert pipette tip down to the tube bottom to remove the colorless bottom layer completely. Therefore only the “fluff” is left in the tube.
- Repeat step 9 and step 10 once.
- Leave the Eppendorf tube cap open to air dry for 5-10 minutes at room temp (do not over dry).
- Add 1× PBS as much as 1-2 volumes of the collected fluff to the tube, and resuspend the “fluff” by pipetting up and downvigorously
- Transfer the suspension carefully into DiagExo® Column (provided) and spin the Column at 2,000× g for 5 minutes to collect all the flow-through;
- The “flow-through” is the isolated pure exosome. Keep it on ice.
- Thaw DiagExo® lysis buffer, Estimate the volume of isolated exosome. Add an equal volume of lysis buffer to the exosome sample for exosomal protein extraction. Pipet up and down the mixture for 5 times till mixed thoroughly. Incubate for 15 minutes on ice.
- Spin at 4°C, 14,000x g for 10 minutes. The supernatant is the extracted exosomal protein. Carefully transfer it to a clean tube, and store at -80°C.
1stSolution A | refer to above table |
2ndSolution B | refer to above table |
3rdSolution C * | refer to above table |
More info: Serum Exosomal Protein Extraction kit
Manual (protocol): Download
Price: 390 Euro
P400 ExoFectin: small RNA into exosome
Storage: store at 4℃
Application: ExoFectin® sRNA-into-Exosome Kit is for the loading of nucleic acids including miRNAs and siRNA into pure exosomes isolated by our kits.
Product Size: 10 reactions
Product Description:
ExoFectin® sRNA-into-Exosome Kit is a proprietary formation designed for the delivery of small RNA including miRNA and siRNA into exosome. This kit provides the following advantages:
Components | Content |
---|---|
ExoFectin Solution A | 700 ul |
ExoFectin Solution B | 400 ul |
Electroporation cuvettes | 10 |
Sterile transfer pipettes | 10 |
The ExoFectin solution is stable for three months at 4°C.
Protocol
- Start with the pure exosomes isolated by our kits (Cat.#: P100, P101, D100 or D101). If the isolated exosome is in the form of pellets, resuspend the exosome pellets in 10-100 ul 1× PBS (depend on the size of exosome pellets, about 10x volume of exosome pellet). Keep the exosome suspension on ice.
- Take 2-5 ul exosome suspension to quantify the exosomal protein. The total exosomal protein concentration represents the quantification of exosome.
- Mix 65 ul ExoFectin Solution A and 35 ul ExoFectin Solution B, combine the mixture with exosomes (10 – 100 ug) and small RNA (0.1 - 3 umol), pipet up and down to mix well. *Total volume of the mixture should not exceed 120 ul.
- Carefully, transfer ExoFectin-exosome-small RNA mixture into a cuvette (the mixture covers the bottom of the cuvette avoiding air bubbles). Cover the cuvette with the cap.
- Insert the cuvette with exosome/small RNA mixture into electroporator cuvette holder. Electroporate the mixture at 400 mV with the pulse time of 10-15 ms.
- Take the cuvette out once the electroporation is completed. Add 500 ul 1× PBS to the cuvette and use the provided “fine tip transfer pipette” to transfer the mixture gently into a new sterile tube preloaded with 1.5 ml 1× PBS containing 1% BSA. Now, the exosomes are loaded with small RNA.
- (Optional) Precipitate the loaded exosomes using PureExo® Exosome Isolation Kit (Cat.#: P100). Refer to P100 manual.
- Downstream application of these loaded exosomes:
- Deliver RNA to target cells with loaded exosomes: resuspend the isolated exosomes in 1× PBS containing 1% BSA. Starve target cells for 48 hours or culture target cells with exosome depleted FBS until cells reach 50% confluence. Apply the small RNA loaded exosomes to the cells. Continue culturing the cells for 48 to 72 hours, and then harvest the treated cells to measure target gene expression using real time RT-PCR.
- In vivo RNA delivery (Intravenous delivery, such as tail injection, or local injection, such as intramuscular injection of loaded exosomes into animals): resuspend isolated exosome in 5% glucose normal saline and inject to recipient animals. Repeated injection will increase the efficiency of the exosome delivery. It is highly recommended to choose the same strain of recipient animal in agree with the exosomes source to minimize the immune rejection. At various time points after the exosome delivery, measure target gene expression in the tissue of interest using real time RT-PCR or detect target gene expression using imaging methods.
Remarks:
The efficiency of electroporation transfection depends on the quality of the exosomes. We do not suggest to start with impure exosome samples prepared by PEG method.
The efficiency of electroporation also depends on the optimal concentration of the exosomes and small RNA. Different RNA and exosome from different source may have different optimal concentration. We suggest customer to test different combinations in the first experiment.
More info: sRNA-into-Exosome Kit (Electro)
Manual (protocol): Download
Price: 245 Euro
P401
Item #: | P401
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Shipping: | Room temperature | |||||||||
Storage: | 4°C (Do not freeze) | |||||||||
Stability: | These reagents are stable for 6 months when store at 4°C. | |||||||||
Application: | ExoFectin sRNA-into-Exosome Kit (chemical) is for the loading of nucleic acids including miRNAs or siRNA into exosomes isolated by our kits. | |||||||||
Product Size: | 100 uL ExoFectin Reagent (~ 20 reactions) | |||||||||
Product Description: | ExoFectin sRNA-into-Exosome Kit (chemical) is a unique blend of polymers designed for the delivery of small RNAs including miRNA and siRNA into exosomes. This transfection reagent provides highly efficient loading abilities of miRNA or siRNA into exosomes, allowing modified exosomes to carry and deliver small RNAs into target cells. This kit provides the following advantages:
» efficient gene delivery
» gentle on treated exosomes
» no small RNA aggregation in exosome
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Kit Contents: |
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Required materials that is not provided in this kit:
- miRNA or siRNA
- Exosome (pure exosome isolated by our PureExo or DiagExo kits is preferred)
- Eppendorf tubes
Positive control (optional): use Alexa Fluor® Red Fluorescent Oligo (Life Technologies, Cat. no. 14750100) to determine transfection efficiency.
More info: ExoFectin (chemical)
Manual (protocol): Download
Price: 245 Euro
P900
Virus-High® AAV / Adv Release Solution
Item #: | P900
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Shipping: | 4°C |
Storage: | -20°C |
Stability: | 1 year in -20℃. |
Application: | fast harvest of Adenovirus (ADV) and Adeno-Associated Virus (AAV) from producing cells, skipping repeated free-thaw steps |
Product Size: | 20 reactions (5 ~ 10 x 106 viral producing cells per reaction) |
Product Description: | Our Virus-High® AAV / Adv Release Solution is for fast and efficient harvest of Adenovirus (ADV), Adeno-Associated Virus (AAV), and other non-envelop and non-budding viruses from producing cells.
» No repeated freeze-thaw steps
» Save 2 to 5 hours (Only need 10 minutes)
» 30% to 60% higher yield of affecting virus (please refer to the following figure for detail.)
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Kit Contents: | AAV / Adv Release Solution 10 mL |
Manual (protocol): Download
Price: 245 Euro
P701-5 Human Wnt-3a
Human Wnt-3a
Purified from human cells by a proprietary process. Xeno-free!
P702-5 Mouse Wnt-3a
Mouse Wnt-3a
Purified from mouse cells
P703-5 Human Wnt-5a
Human Wnt-5a
Purified from human cells by a proprietary process. Xeno-free!
P704-5 Mouse Wnt-5a
Mouse Wnt-5a
Purified from L1 mouse cells
P801 Human Neural Stem Cell
Human Neural Stem Cell (hNSC)
P501 Total Protein Cell
Total Protein Extraction Kit
Name |
3-min Total Protein Extraction Kit (Animal cells) |
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Cat. # |
P501
P501S (Trial size for 5 reactions)
P501L (Large size for 50 reactions)
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Application |
Extract total Protein in as quick as 1 minute. This product is for research use only. | |||||||||||||||
Product Size |
P501: 20 reactions; P501S: 5 rxn; P501L: 50 rxn | |||||||||||||||
Description |
This kit is for extremely fast extraction of total proteins from animal cells (mammalian, insets and other cells). The extraction volume is flexible, from 20 µl to 500 µl, which is useful for limiting number of cells. |
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Shipping / Storage |
Ship at room temperature. Store "Native cell lysis buffer" at 4℃, and the rest at room temperature. (Do not freeze.) | |||||||||||||||
Shelf Life |
12 months | |||||||||||||||
Manual (protocol) |
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Remarks |
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P502 Total Protein Tissue
Total Protein Extraction Kit
Name |
3-min Total Protein Extraction Kit (Animal Tissues) |
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Cat. # |
P502
P502S (Trial size for 5 reactions)
P502L (Large size for 50 reactions)
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Application |
Extract total Protein from animal (human) tissues in as quick as 3 minute. This product is for research use only. | ||||||||||||||||||
Product Size |
P502: 20 reactions; P502S: 5 rxn; P502L: 50 rxn | ||||||||||||||||||
Description |
This kit is for extremely fast extraction of total proteins from animal tissues. The extraction volume is flexible, from 20 µl to 500 µl, which is useful for limiting amount of tissues.
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Shipping / Storage |
Ship at room temperature. Store "Native lysis buffer" at 4℃, and the rest at room temperature. (Do not freeze.) | ||||||||||||||||||
Shelf Life |
12 months | ||||||||||||||||||
Manual (protocol) |
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Component |
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Remarks |
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P503 Plasma Membrane Protein Extraction Kit
Plasma Membrane Protein Extraction Kit
Name |
Plasma Membrane Protein Extraction Kit (mammalian cells or tissues) |
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Cat. # |
P503
P503S (Trial size for 5 reactions)
P503L (Large size for 50 reactions)
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Application |
Rapid extract:
Native total membrane proteins from cultured mammalian cells or tissues (plasma membrane and organelle membrane proteins) Native plasma membrane proteins This product is for research use only. |
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Product Size |
P503: 20 reactions; P503S: 5 rxn; P503L: 50 rxn | |||||||||||||||||||||
Description |
Plasma membrane protein isolation kit is designed to rapidly isolate native total membrane proteins (organelle membrane proteins) and native plasma membrane proteins from cultured mammalian cells or tissues.
This kit is composed of optimized buffers (detergent and EDTA free) and protein extraction filter cartridges with 2.0 ml collection tubes. |
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Shipping / Storage |
Ship at 4℃. Store Buffer A and Buffer B at -20℃ upon arrival, and the rest at room temperature. | |||||||||||||||||||||
Shelf Life |
12 months | |||||||||||||||||||||
Manual (protocol) |
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Remarks |
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P505 Detergent-free Total Protein Extraction Kit
Detergent-free Total Protein Extraction Kit
Name |
Detergent-free Total Protein Extraction Kit (Animal cells) |
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Cat. # |
P505
P505S (Trial size for 5 reactions)
P505L (Large size for 50 reactions)
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Application |
Rapidly extract detergent-free total proteins from cultured cells (mammalian cells, insect cells and other cultured cells) in less than 5 minute. The extracted protein can be used for proteomics (LC/MS), IP, ELISA, 2D-gel analysis, isoelectric focusing, SDS-PAGE, immunoblottings, and other applications. This product is for research use only. |
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Product Size |
P505: 20 reactions; P505S: 5 rxn; P505L: 50 rxn | |||||||||||||||
Description |
This kit is for fast extraction of detergent-free total proteins from animal cells. The extraction does not contain any detergent and EDTA. The extraction volume is flexible, from 20 µl to 500 µl, which is useful for limiting number of cells.
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Shipping / Storage |
Ship at room temperature. Store at 4℃ (Do not freeze.) | |||||||||||||||
Shelf Life |
12 months | |||||||||||||||
Manual (protocol) |
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Component |
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Remarks |
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Our kits optimize your exosome research result.
Cat. #: P101 (10 reactions); P101L (40 reactions); P101S (2 reactions)
Storage: keep all bottles upright, in cool and dark place. Shelf Life: 12 months
Product Size: Each reaction can process 100~500 μL serum or plasma. The yield of each reaction is 100~200 μL exosome, from which 300~400 μg exosomal protein or 200~300 ng exosomal RNA can be extracted.
Product Description (This product is for research use only.)
This kit can isolate / purify pure exosome at high yield from serum or plasma.
Product Contents (store in room temperature)
Component | Amount | ||
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Cat. #: P101 | Cat. #: P101L | Cat. #: P101S | |
Solution A | 2.5 mL | 10 mL | 0.5 mL |
Solution B | 2.5 mL | 10 mL | 0.5 mL |
Solution C | 2.5 mL | 10 mL | 0.5 mL |
Sample Buffer | 20 mL | 80 mL | 4 mL |
PureExo® Column | 10 | 40 | 2 |
* Tightly cap all bottles immediately after each use to prevent evaporation.
Do not process more than 500 μL serum or plasma for each reaction. Otherwise it will cause indistinct layer separation and column clogging.
Protocol (for processing 100-500 μL serum / plasma)
1. Collect the serum or plasma sample and keep it on ice. If start with frozen sample, thaw the sample completely at room temperature, and keep it on ice.
2. Centrifuge the serum / plasma sample at 3,000× g for 15 minutes at 4oC to remove debris.
3. Transfer 100 - 500 μL clear supernatant to a 15mL centrifuge tube without disturbing the pellet. Do not process more than 500 μL sample per reaction. Add Sample Buffer to make a total volume of 2 mL diluted serum / plasma sample, and keep it on ice. (This dilution works well for starting sample range of 100 - 500 μL.)
4. In another 1.5 mL microcentrifuge tube, add Solution A/B/C in the following order to prepare 0.75 mL mixture A/B/C (always prepare mixture A/B/C right before use):
1st add Solution A 0.25mL
2nd add Solution B 0.25mL
3rd add Solution C 0.25mL
* Cap all bottles well immediately after each use to prevent evaporation.
5. Vortex the mixture A/B/C for 10 seconds to obtain a homogenous solution.
6. Add the 0.75 mL mixture A/B/C to the 2 mL diluted serum / plasma sample (from step 3).
7. Cap the 15 mL tube, vigorously vortex for 30 seconds, then incubate at 4°C for 30 minutes.
8. The mixture now appears as 3 layers:
Pipet out the Top transparent layer and discard it without disturbing the Middle fluffy layer.
9. Transfer the Middle fluffy layer (Exosome is in this layer) to another 1.5 mL microcentrifuge tube. Spin the tube at 5,000× g for 3 minutes. A new three-layer separation will appear: Top transparent layer, Middle fluffy layer and Bottom colorless layer. (See figure below.) Proceed to the next step immediately.
Top transparent layer
Middle fluffy layer (Exosome is in this layer.)
Bottom colorless layer
10. Pipet out the Top transparent layer and discard it. Insert pipette tip down to the tube bottom to completely remove the Bottom colorless layer. Only keep the Middle fluffy layer in the tube. Exosome is in this layer.
11. Spin again at 5,000x g for 3 minutes, and 3 layers will appear again. Now, repeat step 10 for one more time. Now only the “fluff pellet” left in the tube. The “fluff pellet” volume is about 25 μL in our case.
12. Leave the tube cap open to air dry for 5-10 minutes at room temp (do not over dry).
13. Add 1× PBS equal to 4 times volumes of the collected fluff pellet to the tube. In our case, we added 100 μL PBS (4 x 25 μL fluff pellet). Resuspend the fluff pellet by pipetting up and down vigorously for 40 times.
14. Shake the tube on a horizontal shaker at high speed for 3 minutes, then pipet up and down vigorously for 10 times. Repeat this “shake-pipet up and down” for another 2 times.
Note: This step is important. If the fluff pellet is not well re-suspended, the exosome may be trapped in the fluff pellet resulting in low exosome purity and yield. Some types of samples, eg. Hyperlipidemia patient serum sample, it is difficult to dissociate the fluff pellet to release exosome. In such case, extend the pipetting and shaking time in step 13 and 14.
15. Spin the tube at 5,000x g for 5 minutes. Without disturbing the “fluff pellet”, transfer the supernatant carefully into one PureExo® Column (provided).
Note: Keep the “fluff pellet” at 4oC. Do not discard it until the experiment is finished. See “Trouble shooting” 2.2 for detail.
16. Spin the PureExo® Column at 1,000× g for 5 minutes to collect all the “flow-through”.
17. The “flow-through” is the isolated pure exosome (exosome suspended in PBS). The whole protocol is completed here. Use it directly for downstream assays (e.g. use 101Bio Exosomal RNA and Protein Extraction Kit, Cat.#: P200, to extract exosomal RNA/Protein), or store at 4°C for up to 1 week , or at ≤80°C for up to 3 months. Concentrated exosome will precipitate after sitting. Pipet up and down to resuspend it well before each use.
More info: AAV - Adv Release Solution (10 mL, 20 Rxn)
Supplier: 101Bio