OBJECTIVES/SPECIFIC AIMS: This research examined 3 aims to address the need to understand and quantify exertion in infants. Aim 1: Develop a schema to identify and code exertional behaviors in infants during treadmill stepping. Aim 2: Establish feasibility for the schema’s use with clinical populations. Aim 3: Pilot the schema in a study designed to induce infant exertion. METHODS/STUDY POPULATION: Aims 1 and 2 were achieved using existing treadmill stepping data. The data used in Aim 1 included eight typically-developing infants (age 7-10 months) who were able to sit independently, but not walk. The data used in Aim 2 came from two separate data sets from infants who took more than 10 steps in a 30-second trial: Data set A included six typically-developing infants (age 2-5 months) who were unable to sit independently (developmentally comparable to atypical populations who might receive treadmill interventions). Data set B included six infants with Spina Bifida (age 3-10 months). Aim 3 was addressed with a prospective study using an exertion model. Pre-walking, typically developing infants (age 8-10 months) underwent five total stepping trials. Trial 1 determined the infant’s individualized maximum stepping speed; trials 2-5 were each 60 seconds and alternated between a baseline stepping speed of.20 m/s and the infant’s maximum stepping speed determined in trial 1. All video data were coded for step type, step frequency, and exertional behavior. RESULTS/ANTICIPATED RESULTS: Aim 1: Two behaviors were identified and determined to capture infant exertion: foot dragging and leg crossing. Aim 2: The feasibility of capturing exertion with these two behaviors was established for young infants and infants with neuromotor delays, with exertional behaviors increasing with stepping exposure (p< 0.05). Aim 3: Total exertion (foot dragging + leg crossing) was higher in the maximum speed trials compared to baseline trials (p = 0.005). DISCUSSION/SIGNIFICANCE OF IMPACT: Exertion in infants can be quantified. The exertion schema developed with this study will support the development of dosing guidelines for infant treadmill intervention. The next step in this line of research is to examine the correlation between infant exertion and heart rate, in effort to move from behaviorally-informed protocols to more precise, individualized protocols based on the physiological response of the infant.