The reconstruction-based (RB) approach can effectively suppress the misdiagnosis problem due to the smearing effect in fault isolation. However, the current exploration of the RB approach for large-scale nonlinear systems is still limited. Therefore, this paper proposes a reliable and effective fault diagnosis method based on a reconstruction-based stacked sparse autoencoder (RBSSAE) for high-dimensional industrial systems. In RBSSAE, a reconstruction-based index achieved by the Steffensen iterative method is developed to check whether the given variable(s) are responsible for the faults efficiently. However, the number of possible faulty variable combinations grows exponentially with the system dimension or actual abnormal variables, causing an unbearable computational burden. Hence, the proposed RBSSAE utilizes a sequential floating forward selection approach to rapidly isolate the most decisive variable combination, meeting a requirement of online fault diagnosis. Finally, the effectiveness of the RBSSAE is verified on a numerical example and a real industrial case.