An experimental methodology is presented to characterize the evolution of structural heterogeneities in cross-linked polymer networks formed by homopolymerizations of multifunctional monomers. In particular, intramolecularly cross-linked macromolecules (ICMS) were synthesized by individual initiation events and characterized. A set of complementary experimental tools, including atomic force microscopy (AFM), size exclusion chromatography, light scattering, and NMR, is used to provide information about size, intramolecular cross-link density, and kinetic chain length for poly(methacrylic anhydride) (PMA) ICMs. Images of PMA ICMs, which contain ca. 1000-2000 monomers, on average, are captured with AFM. Varying polymerization temperature from -13 to 70 degreesC led to an increase in the apparent size of the ICMs and an increase in the fraction of doubly reacted monomers. Furthermore, ICMs synthesized in the presence of tetraethylthiuram disulfide (TED), a photoiniferter precursor, were characterized. Iniferter-mediated polymerization of methacrylic anhydride created smaller ICMs with higher intramolecular cross-link density.