Hello I did an electrophoresis for the first time, and transferred the proteins to PVDF membrane.
I saw the entire ladder in the gel before transferring.
However, after the transfer, half of the ladder disappeared from my gel. I think the transfer was not a complete failure cause I was able to see the entire ladder on the membrane.
Does anyone know the reason behind the disappearance of my ladder (in the gel) after transfer??
Hi Hyun,
May you show an image of both the gel and PVDF? It is possible that some proteins have been completely transferred to the membrane and other partially, due to a wrong or insufficient contact the membrane with the gel in some area?
Your transfer buffer should not have any SDS otherwise there is a chance of over-transfer. Develop the blot and observe you can detect your target protein or not. I have faced the same problem many times. But it did not hamper for detection of my protein. Change your sponge pad. If you use the same sponge pad many times, the thickness of the pad gets reduced which ultimately can cause high current and overtransfer. You can reduce the transfer time but that should not affect your blot. For semidry I use 15volt for 30-35mins and I can detect protein up to 90kDa. Best of luck.
María Luisa Caballero Hello I'm sorry about the delay! These are the pictures of my gel (after transfer and dyeing, the ladder is the fifth band visible) and membrane. As you can see, half the ladder is gone..
Hi Hyun,
Although the entry of the markers in the gel could have been affected by the bubble, it seems that part of the protein loaded of each of the markers have migrated well and have been transferred, as shown the good ladder in the membrane.
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