facilitate pure product with > 90% recoveries in just minutes
product: nanosep
accelerate sample processing
concentrate and purify samples with starting volumes of < 50 µl to 60 ml.
maximize sample recovery
obtain high flow rates and low non-specific protein and nucleic acid binding.
add versatility
available in various membrane types including low-binding bio-inert® (modified nylon), supor® (polyethersulfone), and ghp (polypropylene) membranes, as well as omega™ (modified polyethersulfone) ultrafiltration membrane in a variety of mwcos.
prevent solution bypass
membrane seals stop solution leakage, minimizing sample loss.
easy visual identification
devices are color-coded for a wide variety of membranes, ranging from 1 kd to 0.45 µm.
applications
centrifugal devices can replace traditional separation techniques, such as column chromatography, preparative electrophoresis, alcohol or salt precipitation, dialysis, and gradient centrifugation, when performing the following:
protein or nucleic acid concentration
desalting
buffer exchange
deproteination of biological samples
fractionation of protein mixtures
separation of primers from pcr products
separation of labeled nucleic acids or proteins from unincorporated nucleotides
virus concentration or removal
clarification of cell lysates and tissue homogenates
how to choose the best centrifugal ultrafiltration device
pall’s centrifugal devices simplify many common nucleic acid and protein sample preparation procedures. these devices provide efficient concentration and salt removal of samples from 50 µl to 60 ml in just minutes. choose from membranes that have been developed to assure low non-specific biomolecule binding and typically provide > 90% recovery of target biomolecules.
ultrafiltration method
ultrafiltration is a membrane separation technique used to separate extremely small particles and dissolved molecules in fluids. the primary basis for separation is molecular size, although other factors such as molecule shape and charge can also play a role. molecules larger than the membrane pores will be retained, but not bound, at the surface of the membrane (not in the polymer matrix as they are retained in microporous membranes) and concentrated during the ultrafiltration process.
compared to non-membrane processes (chromatography, dialysis, solvent extraction, or centrifugation), ultrafiltration:
is gentler to the molecules being processed. does not require an organic extraction which may denature labile proteins.
maintains the ionic and ph conditions. is fast and relatively inexpensive.
can be performed at low temperatures (for example, in the cold room).
is very efficient and can simultaneously concentrate and purify molecules.
the retention properties of ultrafiltration membranes are expressed as molecular weight cut-off (mwco) and measured in kilodaltons (kd). this value refers to the approximate molecular weight of a dilute globular solute (i.e., a typical protein) which is 90% retained by the membrane. however, a molecule’s shape can have a direct effect on its retention by a membrane. for example, linear molecules like dna may find their way through pores that will retain a globular species of the same molecular weight.
there are three generic applications for ultrafiltration:
1. concentration. ultrafiltration is a very convenient method for the concentration of dilute protein or dna/rna samples. it is gentle (does not shear dna as large as 100 kb or cause loss of enzymatic activity in proteins) and very efficient (typically > 90% recovery).
2. desalting and buffer exchange (diafiltration). ultrafiltration provides a convenient and efficient way to remove or exchange salts, remove detergents, separate free from bound molecules, remove low molecular weight components, or rapidly change the ionic or ph environment.
3. fractionation. ultrafiltration will not accomplish a sharp separation of two molecules with similar molecular weights. the molecules to be separated should differ by at least one order of magnitude (10x) in size for effective separation. fractionation using ultrafiltration is effective in applications, such as the preparation of protein-free filtrates, the separation of unbound or unincorporated label from dna and protein samples, and the purification of pcr products from synthesis reactions.
membrane selection
based on application these membranes meet the challenges of a wide range of applications with superior performance and stability: omega (modified polyethersulfone) ultrafiltration membrane for rapid concentrating and desalting. bio-inert (modified nylon), supor (polyethersulfone), and ghp (hydrophilic polypropylene) microfiltration membranes for removing particulate (such as gel debris).
choosing the correct mwco
once sample volume is determined, the next step is to select the appropriate mwco (for ultrafiltration) or pore size (for microfiltration). mwcos are nominal ratings based on the ability to retain > 90% of a solute of a known molecular weight (in kilodaltons). table 2 provides retention characteristics of different mwco membranes for some solutes. for proteins, it is recommended that an mwco be selected that is three to six times smaller than the molecular weight of the solute being retained. if flow rate is a consideration, choose a membrane with an mwco at the lower end of this range (3x); if the main concern is retention, choose a tighter membrane (6x). it is important to recognize that retention of a molecule by an ultrafiltration membrane is determined by a variety of factors, among which its molecular weight serves only as a general indicator.
therefore, choosing the appropriate mwco for a specific application requires the consideration of a number of factors including molecular shape, electrical charge, sample concentration, sample composition, and operating conditions.
because different manufacturers use different molecules to define the mwco of their membranes, it is important to perform pilot experiments to verify membrane performance in a particular application.
common variables that increase molecule passage:
sample concentration higher than 1 mg/ml.
buffer conditions that permit molecules to aggregate.
presence of other molecules that increase sample concentration.
lower transmembrane pressure (in the case of centrifugal concentrators, lower g-force).
adsorption to the membrane or device.
low temperature (4 °c versus 24 °c).