Jiangsu Huida Medical Instruments Co.,Ltd

Jiangsu Huida Medical Instruments Co.,Ltd

Precautions For Use Of Ultra-filtration Centrifuge Tubes

(1) Select the appropriate ultrafiltration tube. The molecular weight should not be more than 1/3 of the molecular weight of the target protein, for example, the molecular weight of the target protein is 35kDa, and the ultrafiltration tube with molecular weight can be selected by 10kDa. If the molecular weight of the target protein is about 10kD, it can be used to intercept the molecular weight of 3kD ultrafiltration tube. Read the instructions carefully and pay attention to the different types of ultrafiltration membrane tolerance to various chemicals.

(2) Newly bought ultrafiltration is dry, before use to add ultra-pure water, the volume of the whole membrane, ice bath or refrigerator precooling for a few minutes. Then pour out the water, Centrifuge Tube you can add the protein liquid, add the number, not exceeding the white line on the top of the tube. The operation should be light, before adding the protein liquid, the UF tube needs to be inserted on the ice to precooling.

(3) Balance. Both the quality and the center of gravity must be balanced. Note that the speed and acceleration must not be too fast, or damage the ultrafiltration membrane directly. Centrifuge Tube Start centrifugal ultrafiltration (centrifuge precooling to 4 degrees). The rotational speed RPM of different centrifuges is changed into G, which differs. The acceleration of the centrifuge is tuned to the lowest gear to reduce the pressure on the film. Note, must wait for the centrifuge to achieve the goal speed, the square can leave the centrifuge, otherwise the centrifuge problem, can not deal with the first time. The direction of the membrane and the hinge is adjusted according to the specification (the condition of the angular rotary centrifuge is that the film is perpendicular to the shaft). In the actual use, the general speed of the opening than the instructions in the lower, this can prolong the service life of the centrifuge tube.

(4) When the concentration to the remaining 1ml, take 50ul buffer, add 10ul flow wear, see if there is no change in blue, so as to determine whether the ultrafiltration tube is missing protein. If the tube leaks, pour the upper layer and flow through the new tube and start the ultrafiltration. To accurately determine whether the leakage tube, 5MGML BSA centrifuge 10min, and then take the flow of wear, running protein glue or Bradford Coarse test, continue to add the remainder of the protein liquid concentration (on the ice operation, to prevent the heat of the protein) until all the enrichment liquid is added. During the centrifugal process, pay attention to the occurrence of protein deposition, resulting in blockage. If precipitation occurs, to determine the specific cause of precipitation is the protein concentration is too high or buffer is not appropriate; The former can be solved by ultrafiltration with multiple ultrafiltration tubes, Centrifuge Tube the method of reducing the concentration, the latter is to change the different buffer solution until the protein does not occur precipitate.

(5) The previous steps to concentrate the protein, if you want to change the buffer, when the total protein solution condensed to 1ml or so, gently add the new buffer (after 0.22um ultrafiltration membrane ultrafiltration), and then condensed to 1ml or so, three consecutive times, the final concentration of concentrated final volume depends on the required protein concentration, Generally not more than 500ul, there is also a case of concentration to 200ul. According to the volume concentration of at least 10 times times per time, Centrifuge Tube three times up to 1000 times times, basically can achieve the purpose of buffer.

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