Protein studies play an important role in today’s life science research, especially in fields like proteomics and drug discovery. Protein concentration products are very useful tools for obtaining proteins at the desired concentration while maintaining their characteristics. Vivascience is a leader in this field and provides products for protein concentration based on ultrafiltration and membrane adsorption techniques. They offer several different types of products:
Vivaflow Tangential Flow Filtration
Vivaflow tangential flow filtration systems allow protein concentration and purification for up to 5 liter samples. They consist of a thin channel recirculation path with high cross flow velocities. Protein recovery yield is high and only requires a 10 ml rinse. The unique design also allows the assembly of the tangential flow filtration systems in parallel, allowing for the concentration of 5 liters in less than 75 minutes. Depending on your particular needs, one can select membranes made of different materials, such as polyethersulfone, regenerated cellulose and hydrosart, and various molecular weight cut-offs (MWCO): 5K, 10K, 30K, 50K, 100K and 0.2µm.
Vivacell concentrators are devised for various volumes of proteins, from 20 to 250 ml, and combine the ease of use of centrifugal devices with the flexibility of ultrafiltration cells. The small volume concentrators are very simple although they require extra tubing connections or stirring mechanisms. These devices contain a dead stop that prevents further concentration when residual volume drops below 150µl. The Vivacell 100 for volumes up to 100 ml utilizes pressure, centrifuge, and shaking, while the Vivacell 250, for samples up to 250 ml, can be used on a laboratory shaker for a more rapid concentration.
Vivaspin Centrifugal Concentrators
Vivaspin protein concentrators consist of a vertical membrane design and a channel filtration chamber that provides high-speed concentrations, even with particle-laden solutions. There is a wide range of sizes available for different volumes of samples: Vivaspin 500 for samples up to 500 µl, Vivaspin 2 for sample up to 2 ml, Vivaspin 4 for sample up to 4 ml, Vivaspin 6 for samples up to 6 ml, Vivaspin 15 for 15 ml samples and Vivaspin 20 for samples of 20 ml. There is also a wide array of membranes available such as polyethersulfone, hydrosart, cellulose triacetate, regenerated cellulose. They can be used in either swing out or fixed angle rotors.
Vivascience also has starter kits that contain a selection of MWCOs that one can experiment with while trying to optimize the process. In general, the pore size of the concentrator should be two and a half times smaller than the molecular weight of the protein. However, all concentrators should be tested with a test sample in order to make sure you have selected the right size for your sample.
After choosing the right concentrator, autoclaved distilled water is added to the concentrator which is spun 15 min at 2000 rpm at 4°C to prewash and condition the membrane. Next, the protein of interest is added to the concentrator and spun at the proper speed for about 15-20 minutes. It is advisable to take out a small protein sample and perform a Bradford assay to determine its concentration. If losses are higher than 30%, then the flow-through must be checked and if the protein is detected there, the membrane should be changed or another molecular weight cut off should be chosen.
In some cases, it is better to perform a gradual concentration in order to avoid over concentration and protein precipitation; thus, shorter centrifugation times are advised. If precipitation occurs, either increasing or decreasing NaCl concentration or the addition of detergents or glycerol could help stabilize the protein. Once the concentration volume is reduced and the desired concentration achieved, it is advisable to not add more protein to the concentrator. Instead, the concentrated protein should be removed and a new batch of protein added for a new concentration.
I have used Vivaspin concentrators to concentrate a variety of GST-fused (RanT24N, RCC1, aurora and B, septin2, septin4, septin 5A) and His-tagged recombinant proteins (importin alpha and beta). Susequently, I used the concentrated protein various assays (e.g. immunopulldowns, nuclear import assays). I have also labeled the concentrated protein with rhodamine and injected them into Drosophila embryos and have also used them to generate antibodies in rabbits (aurora A, aurora B, septin2, septin4, septin 5).