Nanoparticles (NPs) are often functionalized with reactive groups such as amines and thiols for the subsequent conjugation of further molecules, e.g., stabilizing polymers, drugs, and proteins for targeting cells or specific diseases. In addition to the quantitative estimation of the reactive conjugation sites, their molecular positioning and nanoscale arrangement on single nanoparticles become more and more important for the tailored engineering and design of functional nanomaterials. Here, we use maleimide or sulfosuccinimidyl ester-modified 1.4 nm gold nanoclusters (AuNCs) to specifically label reactive thiol and amine groups with sub-2-nm precision on metal oxide and polymeric nanostructures. We confirm the binding of AuNCs by measuring and modeling sedimentation properties using analytical centrifugation, imaging their surface distribution and surface distances by transmission electron microscopy (TEM), and comparing the results to ensemble measurements of numbers of reactive surface groups obtained by common photometric assays. We map thiol and amine groups introduced on silica NPs (SiNPs), titania stars (Ti), silica inverse opals (SiOps)., and polystyrene NPs (PS NPs). We show that the method is suitable for mapping local, clustered inhomogeneities of the reactive sites on single SiNPs introduced by masking certain areas during surface functionalization. Mapping precise positions of reactive surface groups is essential to the design and tailored ligation of multifunctional nanomaterials.