A microscale analytical batch chromatographic method for detecting soluble viral DNA-binding proteins in crude extracts.
We describe a micro-method for determining the presence in crude cellular extracts of soluble proteins which can bind to immobilized DNA, using the DNA-binding protein of human adenovirus as an example. Batch chromatography of radiolabeled proteins is performed in microcentrifuge tubes containing 50 microliters packed volume of commercially available denatured calf thymus DNA-cellulose. Eluted single-stranded DNA-binding proteins are then visualized by fluorography following gel electrophoresis. The batch procedure gives yields of adenovirus DNA-binding protein which are comparable to those obtained with a mini-column of similar adsorbent volume. The scale of the procedure makes it convenient for simultaneously analyzing multiple samples.
Department of Biological Sciences, Illinois State University, Normal 61761.
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Khuntirat
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Lucher
D000260:Adenoviruses, Human / Q000276:immunology; D002457:Cell Extracts; D002478:Cells, Cultured; D002482:Cellulose / Q000031:analogs & derivatives; D002845:Chromatography / Q000379:methods*; D004247:DNA; D004268:DNA-Binding Proteins / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015151:Immunoblotting; D011233:Precipitin Tests; D014764:Viral Proteins / Q000032:analysis*
D002457:Cell Extracts; D004268:DNA-Binding Proteins; C033190:DNA-cellulose; D014764:Viral Proteins; D002482:Cellulose; D004247:DNA