Real lab failures, root causes, and fixes — curated and bilingually annotated by our team.
Target protein cannot be reliably detected in whole cell extract despite confirmed expression. Signal may appear in cell culture media but not in cell lysate.
Western blot shows multiple bands instead of a single expected band. Extra bands may appear at different molecular weights than predicted, or background signal throughout the lane is elevated.
Entire membrane appears dark or black after chemiluminescent detection, often with white 'ghost' bands where protein bands should appear. Background signal is saturated across the whole blot.
Proteins smaller than 25-30 kDa show weak or absent signal despite adequate expression levels. Higher molecular weight proteins from the same sample transfer successfully.
Signal intensity is lower than expected or background is elevated. Changing Tween-20 concentration in wash/incubation buffers significantly affects results.
Membrane shows irregular spots, speckles, or splotches distributed across the surface. Background is uneven with localized dark regions rather than uniform signal.
Protein bands appear as vertical smears rather than discrete sharp bands. Smearing may occur above or below the expected molecular weight, making band interpretation difficult.
Target protein bands are absent or very faint on the blot, even after extended exposure times. Positive control samples may also show weak or no signal.
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