Rate of protein transfer from sodium dodecyl sulfate-polyacrylamide gels to nitrocellulose membranes

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About 5μg of protein is generally needed for SDS-PAGE and about 35μg for Western blot analysis. Because purified protein is very precious, we attempted to determine the best transfer efficiency for these analyses that used the smallest amount of protein. We investigated the detection limit of albumin on gel and on nitrocellulose membrane, the transfer rate and time from an electrophoresed gel to a nitrocellulose membrane, and the detection limit of albumin for dot blot analysis and Western blot analysis. We found that the detection limit of albumin was 0.05μg on gel and 0.5μg on nitrocellulose membrane when stained with CBB. The transfer rate from gel to nitrocellulose membrane was 10% and the time was over 90 min. The detection limit of albumin was 0.3μg for the dot blot analysis and 0.5μg for Western blot analysis. Our results suggest that the amount of applied protein can be reduced to almost 1/100th for SDS-PAGE and to between 1/11th and 1/70th for Western blot analysis while still using conventional procedures.

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