Fouling issues are highly undesirable in oil industries, and stable water-oil emulsion is one of the major causes of fouling on pipelines, upgrading equipment, and other surfaces in oil production. Studying the interfacial interactions between emulsion drops and various metal substrates is of significant importance in the fundamental understanding of fouling mechanisms. In this work, surface force measurements using a drop probe atomic force microscope technique and fouling tests were applied to investigate the fouling and antifouling mechanisms of electron-beam-deposited iron substrates with and without electroless nickel-phosphorus (EN) coatings. The effects of oil or aqueous solution conditions have been systematically investigated, including the asphaltene concentration, salinity, pH, and presence of divalent ions. A theoretical model based on the Reynolds lubrication equation and augmented Young-Laplace equation has been applied to analyze the measured force profiles. Our results indicate that the attractive van der Waals force plays an important role in the fouling phenomena, particularly under high-salinity conditions, while the repulsive electric double-layer interaction contributes to the antifouling behavior. Surface force measurements and fouling tests of Fe and EN substrates in toluene-in-water emulsions clearly demonstrate the excellent performance of the EN coating. Our work provides useful insights in the fundamental understanding of fouling/antifouling mechanisms of oil-in-water emulsions on different substrates, with implications to the development of efficient antifouling coatings and strategies in oil production processes.