Preparative high-yield electroelution of proteins after separation by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and its application to analysis of amino acid sequences and to raise antibodies.
A method for the preparative high-yield electroelution of proteins from sodium dodecyl sulphate (SDS) polyacrylamide gel strips was established. The method consisted of SDS-polyacrylamide gel electrophoresis, detection of proteins with sodium acetate and electrophoretic elution at 200 V for 3 h by utilizing a horizontal flat-bed gel electrophoresis apparatus. Standard proteins with molecular masses of 14-66 kilodalton (cytochrome c, aldolase, ovalbumin and bovine serum albumin) were recovered with an average yield of 73.6 +/- 2.3%. A membrane-bound protein, rat skeletal muscle Ca(2+)-ATPase (100 kilodalton) was also well recovered (over 60%). This method was applicable to the purification of proteins required for N-terminal amino acid sequencing and to raise antibodies.
Ohhashi
T
T
Faculty of Pharmaceutical Sciences, Okayama University, Japan.
Moritani
C
C
Andoh
H
H
Satoh
S
S
Ohmori
S
S
Lottspeich
F
F
Ikeda
M
M
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D000252:Calcium-Transporting ATPases / Q000032:analysis / Q000276:immunology; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003574:Cytochrome c Group / Q000032:analysis / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005634:Fructose-Bisphosphate Aldolase / Q000032:analysis / Q000276:immunology; D015151:Immunoblotting; D008969:Molecular Sequence Data; D010047:Ovalbumin / Q000032:analysis / Q000276:immunology; D011506:Proteins / Q000032:analysis* / Q000276:immunology*; D011817:Rabbits; D051381:Rats; D012710:Serum Albumin, Bovine / Q000032:analysis / Q000276:immunology; D012967:Sodium Dodecyl Sulfate
D000906:Antibodies; D003574:Cytochrome c Group; D011506:Proteins; D012710:Serum Albumin, Bovine; D012967:Sodium Dodecyl Sulfate; D010047:Ovalbumin; D005634:Fructose-Bisphosphate Aldolase; D000252:Calcium-Transporting ATPases