Abstract: Quantitative imaging of asialoglycoprotein (ASGP) receptors could estimate the function of the liver. HYNIC-NGA was prepared via introduce bifunctional coupling agent hydrazinonicotinamide to NGA and radiolabeled with ⁹⁹Tc^m by using N,N-bis(2-hydroxyethyl)glycine (bicine) and hydroxyethylethylenediaminetriacetic acid (HEDTA) as coligands, respectively. The labeling yield of ⁹⁹Tc^m-bicine/HYNIC-NGA and ⁹⁹Tc^m-HEDTA/HYNIC-NGA was above 80% under the optimized labeling conditions. After purified with HiTrap desalting column, the radiochemical purity of both ⁹⁹Tc^m-complexes were ＞95%. The biodistribution of ⁹⁹Tc^m-bicine/HYNIC-NGA and ⁹⁹Tc^m-HEDTA/HYNIC-NGA were investigated in normal mice. The blocking experiment of ⁹⁹Tc^m-HEDTA/HYNIC-NGA was performed in normal mice by using GSA (10 mg/kg) as blocking agent. The results of biodistribution showed that the liver uptakes of ⁹⁹Tc^m-HEDTA/HYNIC-NGA were 86.57±5.68%, 74.63±8.74 and 45.11±4.21%ID/g at 5, 30 and 120 min after injection, respectively. While for ⁹⁹Tc^m-bicine/HYNIC-NGA, its liver uptakes were 56.58±6.57, 38.06±3.35 and 23.17±4.96%ID/g, respectively. Both of these tracers showed good initial liver uptake with good retention. The liver uptake of ⁹⁹Tc^m-HEDTA/HYNIC-NGA decreased obviously after blocked by preinjecting free GSA as blocking agent at 5 min after injection, which indicated that the specific could bind to ASGP receptor. Compared with ⁹⁹Tc^m-bicine/HYNIC-NGA, ⁹⁹Tc^m-HEDTA/HYNIC-NGA had higher liver uptake and better retention, and it could be developed as a novel hepatocyte-targeting agent to evaluate hepatic function and clinical use in the future.