In two patients assay of alpha-1,6-amyloglucosidase activity by incorporation of [14C]glucose into glycogen revealed normal activity in leukocytes, erythrocytes, and fibroblasts, whereas no activity was detected in liver and muscle. No activity in any tissue was found when enzyme activity was assayed by following the release of glucose from a phosphorylase limit dextrin. Labeling of glycogen by incubation with crude tissue homogenates according to the protocol used for the [14C]glucose method and subsequent degradation of the outer portion of the polysaccharide molecule with beta-amylase showed that with tissues from normal controls more than 90% of the label of the glycogen was retained in the limit dextrin. When fibroblasts or leukocytes of the patients served as enzyme source up to 80% of the label was released after incubation with beta-amylase or phosphorylase a. Addition of Tris to the assay inhibited enzyme activity in fibroblast homogenates of the patients and of controls to the same extent and had no effect on the distribution of the label between supernatant and limit dextrin after beta-amylolysis of the labeled glycogen. A pH curve performed with fibroblast preparations from the patients and a normal control did not reveal differences in the effect of changes in pH on [14C]glucose incorporation. We propose that incorporation of [14C]glucose into glycogen by the enzyme present in the patients' cells was into alpha-1,4 linkages in glycogen.