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This kit provides a simple and fast 12% SDS-PAGE high-resolution color (green) gel preparation method. This method does not add TEMED, only need AP (ammonium persulfate) to form gel, which greatly reduces TEMED is harmful to the human body and the environment. The gel can be electrophoresed at a constant voltage of 200 V, which can be completed in 30 minutes. Compared with the traditional gel electrophoresis system, this method takes less time and with higher resolution.
This method does not need to add TEMED during preparation, only need AP (ammonium persulfate) to form gel, which greatly reduces the harm of TEMED to the human body and the environment. This product is a modified Bis-Tris formula gel, 25× running buffer is MOPS running buffer, Tris-Glycine running buffer cannot be used. The specific number of gel blocks is related to the thickness of the gel and the size of the gel.
Storage and transportation
Wet ice Transportation ; stored at 4°C and away from light. The BTT electrophoresis buffer (25×) can be stored at room temperature, valid for 12 months.
|BTT upper gel solution A||50mL|
|BTT upper gel solution B (green)||50mL|
|12% BTT lower layer gel solution A||125mL|
|12% BTT lower layer gel solution B||125mL|
|AP (Ammonium Persulfate)||1g (powder)|
|BTT buffer(25 x )||2×250 mL|
|User manual||1 pc|
According to the molecular weight of the target protein, select the appropriate concentration of SDS-PAGE lower gel (separation gel), please refer to the following table:
|AcrylamideConcentration||Best separation range (kDa)|
Prepare 10% AP solution: Get 0.1 g of AP (ammonium persulfate) powder and dissolve it completely with 1 mL of water. The solution is valid for 1-2 weeks when stored at 4°C, and valid for 3 months when stored at -20°C.
According to the experimental requirements, mix solution A and B in equal proportions, add an appropriate amount of 10% AP solution, and prepare the lower gel solution and the upper layer gel solution respectively. For glass plates of different specifications and different thicknesses, the preparation volume of the upper layer gel and the lower layer gel solution can be adjusted in equal proportions. Taking the commonly used 8.3 cm×7.3 cm gel plate as an example, the recommended preparation system as follows:
|Group||Component||Single piece of gel (1.0 mm thickness)||Two pieces of gel (1.0 mm thickness)|
Lower gel solution
|12% BTT lower gel solution A||2.5mL||5mL|
|12% BTT lower gel solution B||2.5mL||5mL|
|10% AP solution||100μL||200μL|
Upper gel solution
|BTT upper gel solution A||1mL||2mL|
|BTT upper gel solution B (green)||1mL||2mL|
|10% AP solution||40μL||80μL|
4.Prepare the gel tool and pour the prepared lower layer gel solution, and immediately add the prepared upper layer gel solution slowly and evenly (distilled water can also be used to seal the lower layer gel liquid surface, 15-30 minutes later until the lower layer gel is fully solidified, then add the prepared upper layer gel solution), insert the comb, and wait for it to solidify (about 15-30 min) before usage.
5.If the prepared gel cannot be used on the same day, it can be stored at 4°C for 1-2 days before usage.
6. Dilute 25 x running buffer to 1 x running buffer before electrophoresis (the running buffer is not recommended to be reused more than 2 times).
7. The whole process of electrophoresis can be 200 V constant voltage electrophoresis, and electrophoresis can be completed in 30 minutes.
1. There is monomer acrylamde in the premix, which is harmful to the human body. Please take protective measures during operation.
2. In order to ensure that the sample enters the separation gel at the same time, when directly pouring the top gel, the prepared top gel solution should be added slowly and evenly.
3. Pour the gel as soon as possible after adding AP, do not leave it for a long time.
4. If you need to further accelerate the gel speed, you can increase the AP usage by 0.5 times.
5. The solidification of the gel has a significant positive correlation with the temperature. Under the same conditions, the higher the temperature, the faster the solidification rate.
6. The company also provides other color (red, yellow, blue)of top gel solutions to distinguish different samples and different gel electrophoresis.
7. The gel made by this kit is not compatible with the Tris-Glycine electrophoresis buffer system, and can only be used with the BTT electrophoresis buffer in the kit. A single product of BTT electrophoresis buffer (25×) is also provided, please refer to G2050.