Electrophoretic extraction of proteins from two-dimensional electrophoresis gel spots

Patent · Sat Dec 31 23:00:00 EST 1988

OSTI ID:866928

Zhang, Jian-Shi ^[1]; Giometti, Carol S ^[2]; Tollaksen, Sandra L ^[3]

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1. Shanghai, CN
2. Glenview, IL
3. Montgomery, IL

After two-dimensional electrophoresis of proteins or the like, resulting in a polyacrylamide gel slab having a pattern of protein gel spots thereon, an individual protein gel spot is cored out from the slab, to form a gel spot core which is placed in an extraction tube, with a dialysis membrane across the lower end of the tube. Replicate gel spots can be cored out from replicate gel slabs and placed in the extraction tube. Molten agarose gel is poured into the extraction tube where the agarose gel hardens to form an immobilizing gel, covering the gel spot cores. The upper end portion of the extraction tube is filled with a volume of buffer solution, and the upper end is closed by another dialysis membrane. Upper and lower bodies of a buffer solution are brought into contact with the upper and lower membranes and are provided with electrodes connected to the positive and negative terminals of a DC power supply, thereby producing an electrical current which flows through the upper membrane, the volume of buffer solution, the agarose, the gel spot cores and the lower membrane. The current causes the proteins to be extracted electrophoretically from the gel spot cores, so that the extracted proteins accumulate and are contained in the space between the agarose gel and the upper membrane. A high percentage extraction of proteins is achieved. The extracted proteins can be removed and subjected to partial digestion by trypsin or the like, followed by two-dimensional electrophoresis, resulting in a gel slab having a pattern of peptide gel spots which can be cored out and subjected to electrophoretic extraction to extract individual peptides.

• Research Organization: Argonne National Laboratory (ANL), Argonne, IL
• DOE Contract Number: W-31109-ENG-38
• Assignee: United States of America as represented by United States (Washington, DC)
• Patent Number(s): US 4824547
• OSTI ID: 866928
• Country of Publication: United States
• Language: English

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