Hi All,
Recently I have trid out Western blotting with ECL detection:
1. The primary antibody is polyclonal, from human and incubated with the
membrane in 100x dilution
2. The secondary anti-human Ab conjugate with HRP in 500x dilution
3. All washes in PBS-T20 for 6x, each time 5 minutes
I got a dark background and clear bands, even the prestained low molecular
weight markers show up as clear bands against the dark background.
Wondering why this happens? I thought the 6 washes should be sufficient.
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