The measurement of microviscosity gradients across different regions of self-assembled systems is crucial for optimizing their performance in both biological and industrial applications. However, this task has long been challenging due to the spatial barrier within the self-assembled systems, the interference from polarity gradients, and the lack of region-positioned probes. To overcome these challenges, we developed three kinds of length-regulated aggregation-induced emission (AIE) probes for the spatial resolution measurement of microviscosity. These AIE probes with varying alkyl chain lengths showed high sensitivity (0.81) over a viscosity range from 2 to 435 mPa s and unaffected luminescence to environmental polarity, ensuring accurate measurements in diverse environments. Using micelles as a model of self-assembled systems, these AIE probes were able to localize selectively in different regions ranging from the hydrophobic core to the hydrophilic shell and interface. The results indicated that microviscosity was highest in the core and gradually decreased toward the outer regions. Furthermore, these AIE probes were successfully applied for monitoring microviscosity in hydrogels and food thickeners, showing a strong correlation among fluorescence intensity, tensile strength, and thickening effects. These findings underscore the potential of length-regulated AIE probes for evaluating the microviscosity in diverse applications.