Market Application & Need
Protein and nucleic acid separation and purification are done routinely in the research laboratory to prepare samples for further analyses. Existing membrane-based solutions cannot separate particles that are close in size and are prone to sample loss. More precise separations require utilizing more complex and expensive chromatography or electrophoresis technologies that are labor intensive and time consuming.
SepCon™ Spin Column Applications and Advantages
Offering the precision of chromatography with the speed and simplicity of a membrane device
- Selective protein separations: separate two proteins that differ in molecular weight by just a factor of two (i.e. cytochrome c and its dimer)
- Separation of ds-DNA fragments: precisely separate DNA oligomers in the 10-300 bp range
- Rapid dialysis: order-of-magnitude faster flow rates – 9X faster than 50 kDa dialysis membrane
- Removal of non-reactive labels: Remove unconjugated reagents from conjugated proteins
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Advantage Example: Rapid Transport and High Permeability
- Objective: Compare the diffusion of small solutes through SiMPore UltraSM® membranes vs. conventional membranes.
- Diffusive transport through SiMPore UltraSM® membrane is almost 10X faster than through similar molecular weight cut-off dialysis membranes.
- Conclusion: Transport through SepCon™ Spin Column is not rate limiting and resulting permeability rates are orders-of-magnitude higher than conventional membranes.
More details are available in the original Nature publication on pnc-Si membranes.
UltraSM® Membranes have almost 10x greater permeability compared to 50kD dialysis membranes
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Advantage Example: Highly Selective Protein Separations
- Objective: Demonstrate size separation precision due to tight pore size distribution of SiMPore’s UltraSM® membranes.
- An UltraSM® membrane separates two chambers in an unstirred diffusion cell. One side contained a solution of 1 mg/ml human serum albumin and 1 mg/ml cytochrome c in phosphate buffered saline (PBS). The second side contained just PBS.
- Three different pore-sized UltraSM® membranes were tested.
- Large pore membrane passed both albumin and cytochrome c. Small size membrane permitted neither to pass.
- Middle pore size membrane with 20 nm cut-off permitted passage of cytochrome c, but not albumin. Furthermore, the membrane was able to retain a dimer of cytochrome c while passing cytochrome c.
- Conclusion: SepCon™ Spin Columns are able to precisely separate proteins that differ in molecular weight by just a factor of two or are different in radius by less than two nanometers.
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Advantage Example: Separation of ds-DNA Fragments
- Objective: Use SiMPore UltraSM® membranes to separate oligomers.
- DNA ladders (10-330 bp) were allowed to equilibrate across an UltraSM® membrane with 15 nm max pore size in a 50 mM NaCl solution.
- Retentate and filtrate were resolved on a gel and the ratio of filtrate/retentate was calculated from densitometry.
- Cut-off occurred between 50 and 60 bp.
- Conclusion: By varying salt concentration and pore size, SepCon™ Spin Column may be used to precisely separate oligomers in the 10-300 bp range.
UltraSM® membranes can be used to separate DNA fragments
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Advantage Example: Removal of unconjugated reagents
- Objective: Use SiMPore UltraSM® membranes to separate free Alexa dye from fluorescently labeled BSA and labeled IgG from BSA.
- Realtime fluorescence microscopy used to monitor passage through membrane.
- Alexa dye passed freely through SiMPore UltraSM® Membrane A (17 nm cut-off) while BSA was almost completely blocked.
- BSA diffuses through SiMPore UltraSM® Membrane B (26 nm cut-off) >4X faster than IgG.
- Conclusion: SepCon™ Spin Column can be used to remove unconjugated reagents from conjugated material and to fractionate proteins that are close in size.
More details are available in the original Nature publication on pnc-Si membranes.
pnc-Si membranes can separate molecules and proteins based on size
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