We explored the potentials of both non-imaging laboratory and airborne imaging spectroscopy to assess arable soil quality indicators. We focused on microbial biomass-C (MBC) and hot water-extractable C (HWEC), complemented by organic carbon (OC) and nitrogen (N) as well-studied spectrally active parameters. The aggregation of different spectral variable selection strategies was used to analyze benefits for reachable estimation accuracies and to explore spectral predictive mechanisms for MBC and HWEC. With selected variables, quantification accuracies improved markedly for MBC (laboratory: RPD = 2.32 instead of 1.33 with full spectra; airborne: 2.35 instead of 1.80) and OC (laboratory: RPD = 3.08 instead of 2.36; airborne: 2.20 instead of 1.94). Patterns of selected variables indicated similarities between HWEC and OC, but significant differences between all other soil variables. This agreed to our results of indirect approaches in which both (i) wet-chemical data of OC and N and (ii) spectra fitted to measured OC and N values were used to estimate MBC and HWEC. Compared to these approaches, we found marked benefits of laboratory and airborne data for a direct spectral quantification of MBC (but not for HWEC). This suggests specificity of spectra for MBC, usable for the determination of this important soil parameter.