Colloidal noble metal nanoparticle aggregates have demonstrated significant advantages in surface-enhanced Raman scattering (SERS) analysis, particularly for online detection, due to their excellent optical properties, spatial homogeneity, and fluidic compatibility. However, conventional chemically induced aggregation methods (such as salt-induced nanoparticle aggregation) suffer from uncontrolled aggregation, limited stability, and narrow detection windows, which restrict their quantitative and long-term applications. In this study, we developed a non-chemical method for fabricating stable colloidal aggregates from uniform β-cyclodextrin-stabilized silver nanoparticles (β-CD@AgNPs) via centrifugation. By precisely controlling the addition rate of silver nitrate, we synthesized β-cyclodextrin-stabilized silver nanoparticles with a uniform size. Surprisingly, these nanoparticles can form highly dispersed and homogeneous colloidal aggregates simply via centrifugation, which is completely different from the behavior of traditional ligand-modified nanoparticles. Notably, the resulting aggregates exhibit excellent SERS enhancement, enabling the sensitive detection of various dyes at nanomolar levels. Furthermore, they maintain a stable SERS signal (RSD = 6.99%) over a detection window exceeding 1 h, markedly improving signal stability and reproducibility compared with salt-induced aggregates. Additionally, using pyocyanin as a model analyte, we evaluated the quantitative performance of these aggregates (LOD = 0.2 nM), achieving satisfactory recovery (82-117%) in spiked samples of drinking water, lake water, and tap water. This study provides a facile strategy for fabricating stable colloidal SERS substrates and paves the way for the advancement of SERS applications in analytical sciences.