To the Editor: We have read the comment by Tilton, Murphy and Mallett (N Engl J Med 287:1100, 1972) regarding the assay of Smith et al. (N Engl J Med 286:583–586, 1972). We have also been preparing the same enzyme and using this assay in a slightly modified form. Our assay employs smaller reaction volumes (100 μl) with a greater volume (75 μl) applied to the phosphocellulose papers (20-mm P-81 paper disk). To reduce the high negative control observed by Tilton et al., we have used a two-minute hot (70 to 80°C) 5 mM Tris wash of the papers, followed . . .