There is now clear evidence that young children's abilities to self-regulate set in motion developmental trajectories that are longitudinally influential, that self-regulation is malleable in childhood with flow-on implications into adulthood, and that efforts to improve self-regulation are enhanced when tailored and adaptive to children's developmental progress. Yet there are significant concerns regarding current approaches to assessing self-regulation, potentially limiting our capacity to respond to children's needs and support their development. In response, this study examined the viability of a novel approach to assessing young children's cognitive, behavioral, and social-emotional self-regulation that uses observer ratings in semi-structured activities. Eighty 3- to 5-year-old children were observed in their preschool in a small group task (a memory card game with other children) and in two individual tasks (an outdoor obstacle course and a curiosity box guessing game). Exploratory factor analysis of resultant Preschool Situational Self-Regulation Toolkit (PRSIST) assessment ratings yielded a two-factor solution which corresponded to cognitive and behavioral self-regulation. Comparison of these observation ratings with task-based (Head–Toes–Knees–Shoulders; HTKS) and adult-report measures of self-regulation (parent and teacher report on the Child Behavior Rating Scale; CBRS) showed strong association with HTKS scores and more modest relations with CBRS ratings. Unlike adult report, however, PRSIST and HTKS were also developmentally sensitive, both showed robust associations with children's school readiness, and neither revealed systematic differences across genders. These findings offer encouraging support for the PRSIST approach, which has high ecological validity and direct relevance to those working with children.