Preparative separation of hemoglobins A and S by gel electrofocusing, using selective zone elution by gel transposition between suitable anolytes and catholytes
| Autor: | Helena Wachslicht, Alice G. McCormick, Andreas Chrambach |
|---|---|
| Rok vydání: | 1978 |
| Předmět: |
Chromatography
biology Elution Isoelectric focusing Hemoglobin Sickle Polyacrylamide Biophysics Hemoglobin A Anemia Sickle Cell Cell Biology Buffers Hydrogen-Ion Concentration Biochemistry Hemoglobins Electrophoresis chemistry.chemical_compound Isoelectric point chemistry biology.protein Humans Hemoglobin Isoelectric Focusing Bovine serum albumin Molecular Biology |
| Zdroj: | Analytical Biochemistry. 85:209-218 |
| ISSN: | 0003-2697 |
| DOI: | 10.1016/0003-2697(78)90291-9 |
| Popis: | Preparative electrofocusing on polyacrylamide gels has been limited, until recently, to excision of gel slices, diffusion, and collection of the slice diffusates. An advance was made by the introduction of a method of selective electrophoretic zone recovery by specific changes of anolyte (A. McCormick, L. E. M. Miles, and A. Chrambach, 1976 , Anal. Biochem.75, 314–324). It was shown (a) that selective zone recovery could be achieved by transposition of the gels into either isoelectric ampholytes or charged buffers, (b) that it could be applied to the gram scale, and (c) that zone elution could proceed either continuously or discontinuously. The early study was, however, limited to a trivial model problem, the separation of hemoglobin from bovine serum albumin (BSA). The present study was an attempt to apply a similar selective zone recovery method to a more demanding separation problem, the separation of hemoglobin A from hemoglobin S as well as from other minor components contained in a sickle-trait human hemolysate. The study shows that selective electrophoretic zone elution from a electrofocusing gel 18 mm in diameter is capable of yielding hemoglobin A, separated from hemoglobin S, differing by only 0.2 pH units in isoelectric point. The recovery of hemoglobin A was 70%, with a load of 32 mg of hemoglobin mixture per gel, using discontinuous zone elution into a collection cup. |
| Databáze: | OpenAIRE |
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