Developing a synthesis-free, multifunctional, and effective colorimetric assay system based on 3,3',5,5'-tetramethylbenzidine (TMB) oxidation is attractive yet challenging. Herein, we established a synergetic Cu 2+ /Cr 6+ -promoting strategy for TMB-based colorimetric detection of Cr 6+ . By introducing Cu 2+ , critical TMB· + ···Cu(I/II)···TMB charge transfer complex (TMC) intermediates were in situ formed to reduce the activation energy of TMB oxidation, thereby accelerating Cr 6+ -mediated TMB oxidation. TMC intermediates also played a pivotal role in H 2 O 2 -participated TMB oxidation, clarifying the secondary responsibility of reactive oxygen species frequently caused by Fenton-like reactions. Leveraging the synergetic capacity between Cu 2+ and Cr 6+ for TMB oxidation, we demonstrated sensitive and specific colorimetric detections for Cr 6+ with a limit of detection of 0.006 μM. With its convenient operation and rapid responsiveness, this strategy successfully enabled the practical detection of Cr 6+ in real water samples. This work not only enhances the understanding of the internal acceleration mechanism in colorimetric sensing but also provides valuable opportunities to advance synthesis-free detection platforms.