In 2023, the U.S. Geological Survey, in cooperation with Riverside Energy Michigan, LLC, conducted an injection experiment to monitor the natural rate of biogenic methane generation in the Mortensen A4-24 well which was completed in the Antrim Shale and located in Antrim County, Michigan, United States of America. Approximately 20 barrels (bbl) of produced water was pumped from the well and stored in a tank in the weeks prior to the injection. On June 6, 2023, ~15 bbl of produced water from the tank was mixed with 2 L of deuterated water (D2O) in a pump truck and injected into the Mortensen A4-24 well. An additional ~5 bbl of produced water from the tank was injected to push the D2O labeled slug into the formation. The well was shut in (both gas and produced water) until August 29, 2023. Produced water and gas samples were collected prior to the injection to assess background conditions and after the shut-in period to assess changes to the system due to the injection. Gas samples were also collected during the shut-in period. This data set includes the geochemical analyses of the produced water samples collected, the compositional and isotopic data of the gas sampled, and metaproteomic data synthesized from extraction of filtered particulates.