OBJECTIVES: Molecular profiling of tumours has become the mainstay of diagnostics for metastasised solid malignancies and guides personalised treatment, especially in nonsmall cell lung cancer (NSCLC). In current practice, it is often challenging to obtain sufficient tumour material for reliable molecular analysis. Cell-free DNA (cfDNA) in blood or other bio-sources could present an alternative approach to obtain genetic information from the tumour. In a retrospective cohort we analysed the added value of cfDNA analysis in pleural effusions for molecular profiling. METHODS: We retrospectively analysed both the supernatant and the cell pellet of 44 pleural effusions sampled from 39 stage IV patients with KRAS (n=23) or EGFR (n=16) mutated tumours to detect the original driver mutation as well as for EGFR T790M resistance mutations. Patients were diagnosed with either NSCLC (n=32), colon carcinoma (n=4), appendiceal carcinoma (n=2) or adenocarcinoma of unknown primary (n=1). Samples collected in the context of routine clinical care were stored at the Netherlands Cancer Institute biobank. We used droplet digital PCR for analysis. RESULTS: The driver mutation could be detected in 36 of the 44 pleural effusions by analysis of both the supernatant (35 out of 44 positive) and the cell pellet (31 out of 44 positive). In seven out of 20 pleural effusions from patients with EGFR mutation-positive tumours, a T790M mutation was detected. All seven supernatants and cell pellets were positive. CONCLUSIONS: cfDNA in pleural effusion can be used to detect driver mutations as well as resistance mechanisms like EGFR T790M in pleural effusion with high accuracy and is therefore a valuable bio-source.