Purpose This study aimed to determine the test-retest reliability and concurrent validity of the International Physical Activity Questionnaire Short Form (IPAQ-SF) in Chilean adults. Methods A cross-sectional validation study was carried out on 161 adults aged between 35 and 65, selected from a population-based study in Temuco, Chile. IPAQ-SF was completed twice, seven days apart, to analyze the test-retest reliability with the intraclass correlation coefficient (ICC). Objective PA was assessed by accelerometry (ActiGraph GT3X+) for seven consecutive days. Intraclass correlation coefficients were used to determine the reliability. Spearman correlation coefficients (rho) and Bland-Altman plots were calculated to assess validity. Results 144 subjects (52.5 ± 8.8 years, 53.9% men) answered the IPAQ-SF on both occasions and had valid accelerometry data. The IPAQ-SF showed moderate reliability for sitting time (ICC = 0.62), while it was poor for walking (ICC = 0.40), moderate PA (ICC = 0.41), vigorous PA (ICC = 0.48), and total PA (ICC = 0.33). There were weak correlations between IPAQ-SF and accelerometry for sedentary behavior (rho = 0.28, p = 0.0005), walking (rho = 0.11, p = 0.17), moderate PA (rho = 0.13, p = 0.128), vigorous PA (rho = 0.18, p = 0.03), and total PA (rho = 0.26, p = 0.002). Conclusions The results suggest that the IPAQ-SF test and retest would provide an acceptable measure of total SB and MVPA, and a weak correlation between IPAQ-SF and accelerometer.