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| Western Blot Forum Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures. |
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#1
08-14-2006, 09:27 PM
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 Western blot troubleshooting Dear Friends Ia m facing a problem with non specific reactivity with proteins in western blott and Iam gettting tooo much background for G6PDH antibody and I need importnat information on the solution used for antibody solutions I am preapraing my primary antibody in 2 percent milk pwder is it right ????? Shall I use PBS to prepare my primary and secondary antibody ????? Please send rely soon Vivek Last edited by admin; 02-24-2007 at 02:56 AM.  |
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#2
08-14-2006, 10:21 PM
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| Re: Western blooot troubleshooting Hey Vivek, welcome. 5 % Blotto is usually standard in 1% TBST. I have never tried PBS although you should be ok. I use 1% TBST for all antibody dilutions. |
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#3
08-15-2006, 05:24 AM
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| Re: Western blooot troubleshooting Dear Let me ask one question can we use PBS to dilute our primary and secondary antibodies rather than TBST and How can I solve the problem of waek signal for my G6PDH protein expression in heart. |
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#4
08-15-2006, 10:41 PM
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| Re: Western blooot troubleshooting Dear Pintu I would load more protein.... or even better, lyse the cells (whole amount) in a small amount of SDS-Page LOADING buffer.. ie 100 ul... this will show your protein for sure... check out [Only registered users see links. ] they have some good articles on this... i think PBS is ok... |
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| blooot , troubleshooting , western |
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