A lipid biomarker study was undertaken to determine the microbial composition and variability in authigenic carbonates and associated soft bottom habitats from the Norfolk and the Baltimore Canyon seep fields along the US mid-Atlantic margin. Results from this study capture a distinct molecular signal from methane oxidizing archaea, including archaeol (I), sn-2-hydroxyarchaeol, pentamethylicosane (PMI), and crocetane. These consortia of methane-oxidizing Archaea have been identified as carrying out anaerobic oxidation of methane (AOM), thereby favoring the precipitation of methane derived authigneic carbonates. The carbon isotope (δ13C) values of AOM-related lipids were strongly depleted in 13C, (i.e., archaeol: -91.64 ‰, sn-2-hydroxyarchaeol: -129.18 ‰, pentamethylicosane (PMI); -131.36 ‰, and crocetane: -70.94 ‰), confirming the dominance of methane as the dominant carbon source for the Archaea during AOM fractionation. The presence of terminally branched fatty acids such as the antesio- and iso-C15:0 components, diagnostic of sulfate-reducing bacteria (SRB), and their depleted δ13C signature (-107.6 ‰), supports syntrophy of SRB with methane-oxidizing archaea. While lipid biomarker profiles of authigenic carbonates biomarker are similar to those found in the seep sediment, suggesting a similar microbial assemblage within the seep-microbiome, a range in lipid composition, distribution, and isotopic signature between seep sites and matrix type suggests AOM is performed by multiple archaeal sources, instead of a single archaeal species.