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A single band on WB smaller than expected


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#1 hepliu

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Posted 13 May 2009 - 12:43 PM

Hi,
I am checking the Mdr expression level in two cancer cell lines by WB using a monoclonal antibody from SCBT. There is a very beautiful band, very specific(?), but the size is about 60KDa, not the predicted 190KDa. If degraded during preparation of cell lysates, it is unlikely that there is only one sharp band on all of the samples (total 8 with different treatments). I called the tech support of SCBT. They gave me a polyclonal antibody to test. There is a prominent band of 60KDa, with many weak (unspecific?) bands. Does anybody have an idea what wrong with my WB?
Any suggestions are appreciated.

Edited by hepliu, 13 May 2009 - 12:51 PM.


#2 bob1

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Posted 13 May 2009 - 05:55 PM

I suggest that the strong band is non-specific, and that you need to expose for a longer time to see your specific band. SCBT have lots of anitbodies but, in my experience, many of them are not very good. I would try another antibody from another company.

#3 hepliu

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Posted 14 May 2009 - 06:15 AM

Thank you for your message.

I have used Li-Cor to expose my membrane. There weren't any other bands except the 60KDa band using the monoclonal antibody. I will try the positive control from SCBT today. Then I will try another antibody from another company.

Edited by hepliu, 14 May 2009 - 06:23 AM.


#4 hepliu

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Posted 14 May 2009 - 06:15 AM

I suggest that the strong band is non-specific, and that you need to expose for a longer time to see your specific band. SCBT have lots of anitbodies but, in my experience, many of them are not very good. I would try another antibody from another company.






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