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Collecting gas-tight fluid samples from deep-sea hydrothermal vents is challenging due to the high pressure, temperature, and corrosiveness of hydrothermal fluids. The most critical part of a gas-tight sampler is the sampling valve because it must have reliable bidirectional sealing capability under such harsh conditions. A new titanium sampling valve has been designed to address these issues. Key features of the new sampling valve include the application of a titanium metal-to-metal seating to achieve high temperature and corrosion resistance, a novel self-tightening design that makes the valve a more reliable closure under high pressure, and a hydraulic actuating mechanism which enhances the convenience and reliability of valve's operation under deep sea. Currently, the sampling valve can be used for vent fluids at depths of 6000 m and temperatures up to 400°C. The new sampling valve has been applied on a gas-tight sampler and tested at the hydrothermal vent sites along the Mid-Atlantic Ridge during the cruise KNOX18RR in 2008. Fluid samples were collected from hydrothermal vents with the depths from 744 to 3622 m and temperatures from 94°C to 370°C. Results of the field tests indicated the usefulness of the new sampling valve to collect deep-sea hydrothermal fluids.