We used long-duration, deep-diving (1000 m) hybrid gliders (Slocum G3, Teledyne Webb Research), equipped with a suite of oceanographic sensors (CTD, fluorometers, and oxygen) and a three-frequency (38 kHz, 67.5 kHz, and 125 kHz) single-beam Acoustic Zooplankton Fish Profiler (AZFP) to conduct biomass assessments of Antarctic krill (Euphausia superba) in the Bransfield Strait and the west shelf of Livingston Island, Antarctica. Two gliders were deployed for 82 and 88 days, respectively, between mid-December 2018 and mid-March 2019. We compare mean biomass density (g m-2) estimates derived from gliders to those from ship-based surveys historically conducted by the U. S. Antarctic Marine Living Resources (AMLR) Program at several temporal and spatial scales to explore the feasibility of using gliders to replace vessel-based biomass surveys. In the Bransfield Strait, glider-based biomass density estimated from 350 glider profiles was 25.0 g m-2, compared with the mean biomass density of 28 g m-2 collected over 13 years of ship surveys. On the west shelf of Livingston Island, glider-based biomass densities estimated from two repeat glider surveys were 61.04 and 104.1 g m-2. Mean biomass density estimates from broad-scale surveys conducted by the U.S. AMLR Program were 55 and 43 g m-2, for January and February surveys conducted between 1997 and 2011. The mean biomass density of krill derived from restricting from the area of ship surveys to overlap with the area surveyed by the glider was 79 g m-2. Mean biomass density estimated from small-scale nearshore surveys conducted by the U.S. AMLR Program from 2005 to 2007, which overlapped spatially with the glider-based surveys, was 82 g m-2. Developing a proper statistical model for the variance estimators from glider-based biomass surveys will require further work. However, these data show that acoustically-equipped glider surveys can provide defensible estimates of krill density and distribution, sufficient to inform management, and should allow the continuation of time series monitoring that has historically been conducted by ships.