and the samples incubated in the dark for 3 hours at room temperature the reduced carboxymethylated samples were analyzed by sds page in this analysis untreated control 2 chain human tissue type plasminogen activator is resolved into three major bands corresponding to type i kringle glycosylation at positions 117 and 184 type ii kringle glycosylation at position 117 and protease lane 1 fig1 endo h digestion of human tissue type plasminogen activator increases the electrophoretic mobility of each kringle band but does not affect the mobility of the protease band lane 2 fig1 the neutral and amino sugar composition of endo h treated t pa is shown in table 2 endo h treatment reduced the mannose content by 5 6 residues per mole and the n acetylglucosamine content by 0 9 residue per mole this result is consistent with stoichiometric removal of the high mannose oligosaccharide from position 117 the fibrinolytic activity of endo h treated human tissue type plasminogen activator was assayed by the in vitro clot lysis assay of collen et al j clin path 21 705 1968 the activity of endo h treated human tissue type plasminogen activator was indistinguishable from that of the untreated control in this assay the human tissue plasminogen samples were iodinated by the iodobead procedure markwell anal biochem 125 427 1982 to a specific activity of approximately 2 u03bcci u03bcg arginine 0 2m and citrate 0 1m ph 6 0 and tween 80 0 01 percent was the buffer used at all times all samples were dialyzed into this buffer prior to iodination ph was adjusted to 8 2 with tris base prior to.