Direct assay of thymidine kinase bound to ion-exchange paper for dot spotting and enzyme blotting analysis.
The direct assay of thymidine kinase (Tk) bound to ion-exchange paper was investigated as a means to further simplify the analytical procedure. Thymidine kinase bound firmly and quantitatively to ion-exchange paper at near neutral pH. The enzymatic properties of Tk did not change while bound to the ion-exchange paper. The amount of phosphorylated 125IdU or 125IdC formed on ion-exchange paper was proportional to the amount of applied Tk. Enzymatic activity could be determined visually by autoradiography or by gamma counting. This method was relatively independent of the protein concentration or volume of the sample and which allows the assay from dilute solutions. A simplified dot spot method that can be used for the assay of thymidine kinase activity in cell extracts is described. Thymidine kinase could also be visualized after electrophoresis and blotting on ion-exchange paper.
D000818:Animals; D001419:Bacteria / Q000201:enzymology; D001853:Bone Marrow / Q000201:enzymology; D004587:Electrophoresis, Agar Gel; D007474:Ion Exchange; D008221:Lymphoid Tissue / Q000201:enzymology; D051379:Mice; D011485:Protein Binding; D013937:Thymidine Kinase / Q000032:analysis*
D013937:Thymidine Kinase