Gold nanoclusters (AuNCs) are promising electrochemiluminescence (ECL) signal probes for their outstanding biocompatibility, unusual molecule-like structures, and versatile optical and electrochemical properties. Nevertheless, their relatively low ECL efficiency and poor stability in aqueous solutions hindered their application in the ECL sensing field. Herein, a facile host-guest recognition strategy was proposed to enhance the ECL efficiency and stability of Au NCs by rigidifying the surface of ligand-stabilized AuNCs via supramolecular self-assembly between cucurbiturils[7] (CB[7]) and l-phenylalanine (l-Phe). Meanwhile, mercaptopropionic acid (MPA) was introduced as a ligand in order to cooperatively enhance the performance of the AuNCs and facilitate the link between AuNCs and bioactive substances. The prepared CB[7]/l-Phe/MPA-AuNCs had a higher ECL emission efficiency, achieving about 2-fold stronger ECL intensity than that of l-Phe/MPA-AuNCs. In addition, after non-covalent modification with CB[7], the finite stability of the papered AuNCs was significantly improved. The prepared CB[7]/l-Phe/MPA-AuNCs showed excellent D-dimer sensing results, exhibiting a linear range from 50.00 fg/mL to 100.0 ng/mL and a detection limit of 29.20 fg/mL (S/N = 3). Our work demonstrated that the host-guest self-assembly strategy provided a universal approach for strengthening the ECL efficiency and stability of nanostructures on an ultra-small scale.