1. Check transfer efficiency by staining gel with Coomassie Blue or the membrane with Ponceau S

2. Optimize time of transfer and current with either a molecular weight marker or a positive control immunoglobulin.

3. Check that gel and membrane make contact during transfer

4. Check that gel and membrane are correctly orientated to the anode of transfer unit

5. Check that excess temperature is not reached during transfer

6. Check that antigenicity is not destroyed by treatment for electrophoresis (SDS, urea, boiling) by dot blotting antigen before and after treatment and detect with ECL reagent