Background: Serum tumor markers are considered a negative prognostic factor in non-small cell lung cancer (NSCLC). Combined detection of serum and pleural fluid (PF) tumor markers is rarely reported. The aim of this study was to analyze the relationship between combined detection of serum and pleural fluid (SPF) tumor markers and prognosis in advanced NSCLC.Results: High levels of SPF CEA (P = 0.001) , SPF CYFRA21-1 (P = 0.001) and SPF CA-125 (P = 0.023) were adversely prognostic factors for overall survival (OS). High levels of SPF CEA (P = 0.003) , SPF CYFRA21-1 (P = 0.001) and SPF NSE (P = 0.019) were related to worse progression-free survival (PFS). In multivariate analysis high levels of SPF CYFRA21-1 was a independent predictor of OS and high levels of SPF CEA and SPF CYFRA21-1 were independent predictors of PFS.Conclusions: High levels of SPF CEA and SPF CYFRA21-1 are correlated with worse survival in advanced NSCLC patients with MPE. The identification of prognostic factors maybe useful in stratifying high-risk populations and will assist the choice of treatment planning.