Uneven transfer / splotchy band pattern (Western Blot)

Question

Accepted Answer

Within a single band the intensity is uneven (light/dark patches); bands across different lanes have inconsistent shapes, with a splotchy appearance. Common causes: 1) Gel-to-membrane contact was poor; transfer pressure was uneven 2) Sample was not fully dissolved — contains particulates or precipitates 3) Antibody solution was not mixed evenly, or antibody aggregated 4) Transfer apparatus has unstable contact or current Solutions: 1) Reassemble the transfer sandwich carefully; ensure gel-membrane contact is flat (no bubbles) 2) Centrifuge samples before loading; if needed, filter (0.22 µm) 3) Prepare antibody fresh and mix thoroughly before applying 4) Check transfer buffer composition, current setting, and consumable quality