We examined the sensitivity of GRACE Follow-On (GRACE-FO) laser ranging interferometer (LRI) measurements to sub-monthly time-variable gravity (TVG) signals caused by transient, high-frequency mass changes in the Earth system. GRACE-FO LRI provides complementary inter-satellite ranging measurements with higher precision over a wider range of frequencies than the baseline K-band microwave ranging system. The common approach for studying mass variation relies on the inverted TVG or mascon solutions over a period of, for example, one month or 10 days which are adversely affected by temporal aliasing and/or smoothing. In this article, we present the alternative along-orbit analysis methodology in terms of line-of-sight gravity difference (LGD) to fully exploit the higher precision LRI measurements for examination of sub-monthly mass changes. The discrepancy between “instantaneous” LGD LRI observations and monthlymean LGD (from Level-2 data) at satellite altitude indicates the sub-monthly gravitational variability not captured by monthly-mean solutions. In conjunction with the satellite ocean altimetry observations, highfrequency non-tidal atmosphere and ocean models, and hydrology models, we show that the LGD LRI observations detect the high-frequency oceanic mass variability in the Argentine Basin and the Gulf of Carpentaria, and sub-monthly variations in surface (river) water in the Amazon Basin. We demonstrate the benefits gained from repeat ground track analysis of GRACE-FO LRI data in the case of the Amazon surface water flow. The along-orbit analysis methodology based on LGD LRI time series presented here is especially suitable for quantifying temporal and spatial evolution of extreme, rapidly changing mass variations. Plain Language Summary The GRACE Follow-On (GRACE-FO) mission, launched on 22 May 2018, is extending the legacy of GRACE by measuring time-variable gravity field of the Earth. Temporal variation in Earth gravity field is caused by mass change on and near the Earth surface due to various processes in hydrosphere, cryosphere, solid Earth, and ocean. In addition to the Ka-band ranging system, the GRACE-FO satellites are also equipped with a laser ranging interferometer (LRI) which measures the inter-satellite distance change with higher precision. When GRACE-FO satellites fly over a mass change anomaly, the LRI measurements capture the corresponding gravitational changes at ∼500 km altitude. We directly examine the LRI gravitational measurements and show that they clearly detect the rapid mass variability associated with the high-frequency (a) oceanic gyre in the Argentine Basin, (b) oceanic mass changes in the Gulf of Carpentaria, and (c) river water variation in the Amazon Basin. Such processes show large mass variability within a month and, thus, they cannot be adequately studied using the standard monthly-mean data products from GRACE-FO mission. Therefore, the presented along-orbit analysis approach broadens the scope of geophysical applications that can be addressed by GRACE-FO satellites data.