APPLICATION OF HIGH-AFFINITY BINDING CONCEPT TO RADIOLABEL AVIDIN WITH TC-99M LABELED BIOTIN AND THE EFFECT OF PI ON BIODISTRIBUTION

In order to label avidin with Tc-99m, we took advantage of the high affinity binding of biotin to avidin; we radiolabeled a biotin derivative with Tc-99m and then bound this Tc-99m labeled biotin derivative to avidin. For our labeling approach, N epsilon-biotinyl-L-lysine (Biocytin) was reacted with the N-hydroxy-succinimide ester of benzoylmercaptoacetyltriglycine (Bz-MAG3). The resulting Bz-MAG3-Biocytin was labeled with Tc-99m using Tc-99m glucarate as a Tc-99m transchelating agent and mixed with avidin at a 1:1 molar ratio resulting in almost a quantitative labeling yield. Tc-99m-MAG3-Biocytin/Avidin was stable in serum at 37 degrees C with 97 and 95% of the total Tc-99m activity still bound to avidin at 2 and 24 h, respectively. The biodistribution of Tc-99m-MAG3-Bio-cytin/Avidin in normal Balb/c mice showed a high liver and kidney uptake with 56.6 and 28.9%, respectively at 10 min. We attempted to lower the liver and the kidney activities by reducing the isoelectric point (pI) of avidin by conjugating succinic acid moieties at lysine residues of avidin (pI 10). The kidney uptake decreased to 19.0, 3.1 and 1.7% when the pI of avidin was reduced to 7.0-9.3, 5.5-6.2 and 4.0-4.8, respectively. The lowering of the pI, however, did not change the liver activity appreciably.