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Application of Buffer Bicine, Tris, MOPS in the Experiment of Protein Transfer
Bicine, Tris, and morpholine propane sulfonic acid MOPS are all commonly used buffers in biology, mainly used for protein and nucleic acid related research experiments. In some experiments, not only a single buffer is used, but a combination of several.
In the protein polyacrylamide transfer membrane experiment, the loading buffer uses the Tris buffer, the running buffer uses the Tris-MOPS-SDS system, and the transfer buffer uses the Tris-Bicine system, which can separate the molecular weight between 205-45KDa The electrophoresis is fast in about 40 minutes, and the transfer efficiency is high in 30 minutes to complete the transfer of all proteins.
Desheng Bicine, Tris, MOPS raw material manufacturers supply
5X Sample Buffer (5 times working concentration sample buffer):
SDS 1.0 g, Glycerol 5.0 ml, bromophenol blue 25 mg, Tris base 150 mg (adjust the pH to 6.8), 2-mercaptoethanol 1.0 ml, and deionized water 10 mL.
10X Running Buffer:
Tris base 60.6g, morpholine propanesulfonic acid MOPS 104.6g, EDTA3.0g, SDS 10.0g, deionized water 1 liter.
Transfer Buffer 20X Transfer Buffer:
Tris base 60g, Bicine 81.6g, deionized water 1 liter.
When performing electrophoresis in the electrophoresis tank, put the electrophoresis gel in the tank and pour enough 1X electrophoresis buffer so that the buffer covers the loading gel hole by 5-7mm. Pour enough 1X electrophoresis buffer into the outer tank to ensure the cooling effect during electrophoresis. It is best that the buffer level of the outer tank is close to the bottom of the sample hole. Using a pipette or syringe, flush the glue hole to remove air bubbles and buffers present in the glue hole. The maximum sample amount of protein per well is 50ug (the sample is incubated in 1X sample buffer at 100°C for 5 minutes). The best sample amount must be obtained through experiments. Excessive sample loading will cause smear or band distortion. Excessive free carbohydrates in the sample can also cause band distortion. 140V for 30-45 minutes, the specific time depends on the size of the target protein.
After the electrophoresis is over, take out the electrophoresis gel according to the equipment instructions, and perform staining or transfer experiments. Take 50 mL of 20X transfer buffer, use 100 mL of methanol or ethanol and 850 mL of deionized water to prepare 1X transfer buffer. Wet transfer membrane conditions are 40V, 90 minutes; or semi-dry transfer membrane conditions are 25V, 30 minutes.
In the protein gel electrophoresis transfer experiment, three biological buffers, Bicine, Tris, and MOPS, are used. This experiment requires a certain degree of operating experience, and novices can try more. In addition to the buffer solution, Desheng also has a variety of buffer materials, which can be configured into different buffer systems according to needs, which is more economical and flexible for experiments.