Background: Flux variability analysis is often used to determine robustness of metabolic models in various simulation conditions. However, its use has been somehow limited by the long computation time compared to other constraint-based modeling methods. Results: We present an open source implementation of flux variability analysis called fastFVA. This efficient implementation makes large-scale flux variability analysis feasible and tractable allowing more complex biological questions regarding network flexibility and robustness to be addressed. Conclusions: Networks involving thousands of biochemical reactions can be analyzed within seconds, greatly expanding the utility of flux variability analysis in systems biology. Background Flux balance analysis (FBA) [1,2] is concerned with the following linear program (LP) max v c v Sv 0 v v v T l u subject to = (1) where the matrix S is an m