The operation of concentrating the centrifuge tube is simple. Select the appropriate concentrated centrifuge tube, add the sample to be treated, perform centrifugation according to the instructions of the centrifuge tube, and finally recover the intercepted sample. Under the action of centrifugal force, the small molecule solute and solvent in the solution pass through the ultrafiltration membrane and are collected in the filtrate collection bottle, while the macromolecular solute is trapped in the concentrated centrifuge tube by the ultrafiltration membrane. The main basis for this separation is the size of the molecule, but other factors such as molecular shape and charge can also be affected.
In general, the recovery rate of ultrafiltration can reach more than 90%. Most of the sample loss is due to non-specific adsorption of the membrane surface and the surface of the vessel. Therefore, it is very important to choose a high-quality concentrated centrifuge tube with good reputation.
How to choose a concentrated centrifuge tube
Different series of concentrated centrifuge tubes are suitable for processing different volumes of samples. You can choose according to your sample size. However, if the sample concentration is low and the volume is large, the smaller volume of the concentrated centrifuge tube can be selected to repeat the sample centrifugation.
2. Choose the appropriate molecular weight cut off (MWCO)
Once the sample volume is determined, the next step is to select the appropriate MWCO. For proteins, it is recommended that the MWCO of the selected membrane be 1/6-1/3 of the size of the molecule to be retained. Generally, the smaller the cut-off aperture, the slower the flow rate, but the greater the rejection ratio. Therefore, if the flow rate is the main consideration, it is recommended to choose a filter with MWCO of 1/3; if the concentration ratio is considered, it is better to choose a denser membrane.
Application of concentrated centrifuge tubes
This is an application that everyone is familiar with. Ultrafiltration facilitates the concentration of diluted protein or DNA/RNA samples. It is gentle, does not interrupt up to 100 kb of DNA, does not cause loss of protease activity, and is also very efficient, with recovery rates typically above 90%.
Desalting and buffer exchange
Ultrafiltration brings a convenient and efficient method to remove and exchange salts in solution, remove denaturing agents, separate complex molecules, remove low molecular weight components, or rapidly change ion or pH environments. For example, when digesting with two or more restriction enzymes, a common digestion buffer may not be found, and the buffer may be replaced by a concentrated centrifuge tube.
Of course, ultrafiltration does not completely separate two molecules with close molecular weights. For efficient separation, the molecular weight of the molecules to be separated must differ by at least an order of magnitude (10X). For the purification and recovery of PCR products, ultrafiltration is suitable. The PCR reaction mixture contains a variety of salts, free nucleic acids, glycerol, proteins and primers, so most downstream applications require some purification of the PCR product. The ultrafiltration method is fast, requires little operation, high yield, and does not damage DNA.