The first stage in microcolony formation and biofilm development is the attachment of bacteria to the surfaces, involving the aggregation of microbes beneath a self-assembled structure of extracellular polymeric substances (EPS). Exopolysaccharides, different kinds of structural proteins, lipids, enzymes and extracellular DNA make up the bulk of this organized structure. It imparts rigidity and also serves as an external organ for digestion. A crucial regulatory compound, cyclic-di-GMP modulates its activities and promotes the shift from the free planktonic stage to biofilm-forming sessile stage. The EPS of Enterobacteriaceae primarily consists of curli amyloid fibres. Disruption of EPS structural components with enzymes such as glycoside hydrolase and DNase is possible for bacterial biofilm. Microorganisms living inside biofilms are significantly more resistant to antibiotics when compared to motile free-living microbes and this resistance has become a threat to human specially in the clinical environments as well as food industries, regardless of whether there are strict protocols to mitigate their development and propagation. For better observation and understanding of the intricate architecture of biofilm, modern methods of imaging SEM, VPSEM and FIBSEM have been proven very beneficial. Advanced research on the architectural framework of biofilm and advanced techniques for its destruction remains an important concern for human welfare.