Weird Western Blot (circular streaks)
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Weird Western Blot (circular streaks)
I've had a couple of weird western blots lately,
we use a standard SDS PAGE tris glycine minigel
(BIORAD), and transfer using the minigel
apparatus at 100v for 90 minutes with a cooling
block (ice in a container) present. The transfer
buffer is the usual methanol/glycine/tris recipe.
The problem shows up when ponceau red stained as
weird circular streaks over some (not all) of the
blot. Coomassie staining the gel reveals ... ...
Last update 02-Apr-2004, Rating Fair of 1 votes.
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I am looking for anti-mouse collagen I and collagen IV antibodies for western blotting. The ones we have in our lab work only in immunohistochemestry. I have searched in eweb site of several compagnies, but didnt found anything interesting. It will be very helpful in somebody can provide me with adequate information.
Thank you very much
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We have been doing western blotting for collagen (RTT),the secondary Ab being conjugated with HRP.We see white bands after developing with DAB against brown background. We use skim-milk powder for blocking.Can anybody help us trouble shoot this problem?
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we see our blots (collagen samples) very distinct in ponceau stain.but dont get the bands after western blotting,the membrane turns brown after adding the developer which we use is DAB.the enzyme we use is HRP.we dont kno were we go wrong.our antibody concentration or the steps in western blotting.we keep for blocking one hour.can some one say the dilution factor for antibody of the type I collagen ,our collagen is RTT.We need ur help very urgent.
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