Soil organic carbon (SOC) is a primary constituent of soil organic matter and plays an important role in the regulation of many soil processes, including greenhouse gas emissions. Recently, SOC also became an indicator for monitoring climate change mitigation policies in the agricultural sector. The availability of up-to-date SOC inventories is thus crucial in terms of supporting SOC–related actions at country or sub-country scales. Currently, the National Monitoring System of the Agricultural Soils of Slovakia (CMS-P), whose network of 318 monitoring sites was last surveyed in 2018, is the only available source of up-to-date topsoil SOC data for agricultural land in Slovakia. Although very useful at the national scale, the number of CMS-P observations it contains is too limited for much needed sub-national SOC inventories. We hypothesized that with the aid of well-chosen macro-scale drivers of topsoil SOC accumulation in agricultural land in Slovakia, and by mapping those drivers geographically, we could upscale the CMS-P observations and produce a regional estimate of topsoil SOC. Altitude, land cover, topsoil texture, and soil type were assumed to be the key factors controlling topsoil SOC accumulation in Slovakia, and based on these, the country was classified into 14 macro-scale geographical regions. Typical ranges and mid-class values of 0–30cm topsoil SOC concentrations (%) and stocks (t ha−1) were calculated for each macro-scale region from CMS-P data. The average topsoil SOC content in agricultural land was estimated to be 2.13% (72.9 t ha−1). The highest topsoil SOC stock (> 90 t ha−1) was estimated for the lowlands of Slovakia, and the lowest (< 50 t ha−1) for the shallow and stony soils of mountain regions. When aggregated to 78 administrative regions at LAU1 level, the area-weighted averages ranged between 39.20 t ha−1 and 80.0 t ha−1, with the highest values (> 65 t ha−1) being in LAU1 regions in the south-west, south-east, and north of Slovakia where arable land is most prevalent. Total SOC storage in 0–30cm topsoil of agricultural land in Slovakia was estimated at 118.39 Mt, with two-thirds of this amount stored in arable soils in 33 south-west, south-east, and south LAU1 administrative regions. As there is no alternative and up-to-date dataset on topsoil SOC content in Slovakia, the upscaling algorithm presented in this study is an important step toward utilizing CMS-P data for sub-national SOC inventories. It may also offer a new way of providing inputs to help predict future or alternative regional topsoil SOC accumulation trajectories in Slovakian agricultural land using process-based or statistical models.