Mapping the longitudinal relaxation time (T 1 ) of brain tissue is of great interest for both clinical research and MRI sequence development. For an unambiguous interpretation of in vivo variations in T 1 images, it is important to understand the degree of variability that is associated with the quantitative T 1 parameter. This paper presents a general framework for estimating the uncertainty in quantitative T 1 mapping by combining a slice-shifted multi-slice inversion recovery EPI technique with the statistical wild-bootstrap approach. Both simulations and experimental analyses were performed to validate this novel approach and to evaluate the estimated T 1 uncertainty in several brain regions across four healthy volunteers. By estimating the T 1 uncertainty, it is shown that the variation in T 1 within anatomic regions for similar tissue types is larger than the uncertainty in the measurement. This indicates that heterogeneity of the inspected tissue and/or partial volume effects can be the main determinants for the observed variability in the estimated T 1 values. The proposed approach to estimate T 1 and its uncertainty without the need for repeated measurements may also prove to be useful for calculating effect sizes that are deemed significant when comparing group differences.