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Affinity chromatography of reticulocyte lysates on haptoglobin improves two‐dimensional mapping of translation products
Author(s) -
Rademacher Bruce E.,
Steele William J.
Publication year - 1987
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150081106
Subject(s) - reticulocyte , translation (biology) , chromatography , haptoglobin , hemoglobin , elution , chemistry , affinity chromatography , sodium dodecyl sulfate , resolution (logic) , biochemistry , biology , messenger rna , enzyme , artificial intelligence , computer science , immunology , gene
Abstract A simple method is described for enhancing the resolution and detectability of translation products in two‐dimensional mapping of reticulocyte lysate translation systems. Assays are fractionated by affinity chromatography on human haptoglobin‐Affi‐Gel 15 to remove hemoglobin and translation products are eluted with sodium dodecyl sulfate, buffered at pH 9.2. Translation products purified in this, way have less than 10 % of the hemoglobin present in the original assay. The recovery of translation products is about 95 % after haptoglobin chromatography and about 75 % after concentration for mapping. Hemoglobin removal not only enhances the resolution and detectability of low and moderate abundance translation products but also reduces horizontal and vertical streaking in two‐dimensional mapping without significantly altering the distribution of translation products.