The U.S. Naval Oceanographic Office used Deepstar 4000 for 13 dives during October and November 1967. Marine geology, biology and the physical properties of the water column were studied on the 10 deep dives of this series. These dives were accomplished along the east coast of the United States and in the Caribbean. Great similarities in the bottom features at widely separated sites as well as dissimilarities in adjacent areas were particularly noteworthy.

During this operation Deepstar 4000 was evaluated as a Deep Oceanographic Survey Vehicle (DOSV). The lack of an all-weather capability and the rather limited payload hampered this study, but the overlapping fields of the viewports and the ability to operate in very close proximity to the bottom, regardless of terrain, are desirable features that should be included on any future DOSV.^[3][4]

Accurate measurements of in-situ sound speed, temperature, salinity, and pressure have been achieved during numerous replicating dives aboard Deepstar 4000 to depths of 1,200 metres (3,900 ft). Crewed deep submergence vehicles offer optimum methods for observing the actual ocean environment. Advantages of submersibles include: capabilities of mounting multiple equipments with short cables, visually monitoring instruments during data acquisition, and controlling proximity to the seafloor—significant improvements over suspending sensors with miles of cables from rolling, pitching surface craft.

On one Deep Star 4000 dive south of San Diego, California, the crew, Dr. Eugene C. La Fond and pilot, narrowly escaped tragedy when the ascent system and its backup failed at 3,500 feet down. The weights normally meant to detach to allow ascent would not release. To save the craft, hundreds of pounds of mercury ballast used for trim was hand pumped onto the ocean floor and the craft could rise.^[5]

Instrument packages have consisted of three precision velocimeters (two NUS TR-4's and one TR-5), two Dymec temperature sensors, one Bissett-Berman salinometer, one or two Vibrotrons, and four Fjarlie bottles with four reversing thermometers each. Accuracy is enhanced by meticulous calibrations before and after dive series, delicate handling of all equipment, intercomparison of several instruments, consistently careful measurements, and correct assessments of thermal lags and pressure effects. Results are compared with existing equations for sound speed versus temperature, salinity, and pressure. The United States has made a torpedo-shaped probe called the Deep Flight.^[3]

In 1967 a crew member of the Deepstar 4000, Joe Thompson, claimed that he had observed a fish of extreme size while onboard the vessel. According to his account the fish was observed in 1966 by himself and other crew members while off the coast of San Diego, he claimed to have observed the fish while placing equipment on the sea floor and glancing outside his window. His description of the creature was that it was bigger than the submarine itself and that it's eyes were larger than dinner plates. According to Thompson the creature quickly swam away before anyone could get photographic evidence of it.^[6] Following this account Thompson would continue to spread this story to others, such as Cryptozoologist Gardner Soule.^[7]