This work presents a probabilistic method to assess the seismic vulnerability of historical masonry structures, such as churches or cathedrals, including uncertainty analysis of the material parameters. The proposed approach considers the pushover analysis using the finite element method for the structural evaluation of the seismic behaviour of masonry structures. A stochastic analysis based on Monte Carlo simulation investigates the effect of the uncertainty of the structural members’ mechanical parameters on the evaluation of the seismic fragility. The method is applied to the seismic assessment of the bay structure of Santa Maria del Mar church in Barcelona, Spain. This case study is characterised by complex geometry and material heterogeneity, and shows to be sensitive enough to the uncertainty of the material properties to experience two possible collapse mechanisms in case of an earthquake. The study presents how to derive analytical seismic fragility curves by considering the uncertainties regarding the material properties and the different types of collapse mechanism.