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RATIONALE: The recent development of compound-specific online chlorine isotope analysis ((37) Cl-CSIA) methods has fostered dual chlorine-carbon isotope studies to gain better insights into sources and environmental transformation reactions of chlorinated ethenes (CEs). 1-point and 2-point calibration schemes are currently used to convert raw data to the international δ(37) ClSMOC scale, but a critical evaluation of best practices to arrive at reliable δ(37) ClSMOC signatures and enrichment factors was missing and is presented here. METHODS: Aqueous solutions of neat PCE and TCE and aqueous samples from a TCE biodegradation experiment with pure cultures of Desulfitobacterium hafniense Y51 were analysed for their chlorine isotope ratios using GC/qMS and GC/IRMS. The δ(37) ClSMOC values were obtained using 1-point and 2-point calibration schemes. Chlorine isotope enrichment factors, ɛCl , were calculated using both approaches and the corresponding bias of δ(37) ClSMOC values introduced by the different types of calibration was determined. RESULTS: Different calibration methods resulted in significant differences (up to 30%) in both δ(37) Cl signatures and ɛCl -values. CONCLUSIONS: Our results demonstrate that a 2-point calibration together with comprehensive information on reference materials is indispensable and should become standard practice for reliable (37) Cl-CSIA of organic compounds.