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Researchers have yet to apply a formal operationalized theory of motivation to neurobiology that would more accurately and precisely define neural activity underlying motivation. We overcome this challenge with the novel application of the Expectancy Theory of Motivation to human fMRI to identify brain activity that explicitly reflects motivation. Expectancy Theory quantitatively describes how individual constructs determine motivation by defining motivation force as the product of three variables: expectancy – belief that effort will better performance; instrumentality – belief that successful performance leads to particular outcome, and valence – outcome desirability. Here, we manipulated information conveyed by reward-predicting cues such that relative cue-evoked activity patterns could be statistically mapped to individual Expectancy Theory variables. The variable associated with activity in any voxel is only reported if it replicated between two groups of healthy participants. We found signals in midbrain, ventral striatum, sensorimotor cortex, and visual cortex that specifically map to motivation itself, rather than other factors. This is important because, for the first time, it empirically clarifies approach motivation neural signals during reward anticipation. It also highlights the effectiveness of the application of Expectancy Theory to neurobiology to more precisely and accurately probe motivation neural correlates than has been achievable previously.Expectancy Theory provides a formal operational definition of motivation.For the first time, we apply Expectancy Theory to fMRI to study motivation signals precisely.With Expectancy Theory, motivation fMRI signals are isolated from reward and other processes.Signals in ventral striatum, midbrain, sensorimotor, and visual cortex scale specifically with evoked motivation.This highlights the effectiveness of applying Expectancy Theory to probe motivation neurobiology.