To conjugate the multifunctional dendrimer onto the surface of the ACC-based cores, 20 mg of ACC@DOX.Fe2+-CaSi was first dispersed into anhydrous toluene and reacted with 10 μl of (3-aminopropyl) trimethoxysilane at 60°C for 24 hours under reflux. The reaction lasted for 24 hours, and the nanoparticle product was washed twice with ethanol. Subsequently, 20 mg of ACC@DOX.Fe2+-CaSi-NH2 was dispersed in 10 ml of ethanol and reacted with 5 mg of TOS-PEG4000-N3 at 45°C overnight under stirring to obtain ACC@DOX.Fe2+-CaSi-N3. The multifunctional dendrimers were eventually immobilized onto the nanoparticle surface via click reaction. Specifically, 20 mg of ACC@DOX.Fe2+-CaSi-PEG-N3, 20 mg of mPEG-GPLGVRGDGG-PAMAM, 2.1 mg of PAMAM-FA, 2 mg of pentahydrate copper sulfate, and 4 mg of ascorbic acid were mixed in 5 ml of DMF and stirred at room temperature for 24 hours under a nitrogen atmosphere. The nanoparticles were collected via centrifugation and washed twice with ethanol. The purified product was also stored in ethanol to prevent degradation.

Some of the nanopreparations were also labeled with FITC for supportive characterizations. The synthesis protocol of these samples was almost identical to that of ACC@DOX.Fe2+-CaSi-PAMAM-FA/mPEG, the only difference is that 4 mg of FITC was added for the co-condensation with ACC instead of the DOX·HCl and anhydrous ferrous chloride.

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