The motility of microorganisms such as Escherichia coli (E. coli) in complex biological fluids is a topic of significant interest in medicine. Most biological fluids, including mucus, exhibit complex rheological properties such as yield-stress and viscoelasticity. Yield-stress fluids behave as solids under low stress and flow as liquids under high stress, presenting unique challenges and opportunities for microbial motility studies. However, studying motility in yield-stress fluids systematically is challenging as reproducing biological fluids with similar rheological properties is difficult. This thesis provides a framework to investigate E. coli motility in complex fluids using aqueous solutions of tethered hairy nanoparticles (HNPs) as a potential model system. To ensure the suitability of HNPs for this purpose, their biocompatibility with E. coli was evaluated using the disk-diffusion and spectrometer growth rate methods. Next, Differential Dynamic Microscopy (DDM), a technique based on the principles of Dynamic Light Scattering, was employed to quantify the motility parameters of high-density E. coli suspensions, and the accuracy of this method was verified. Following this, a protocol for culturing motile E. coli was developed, along with a framework for capturing their movement with an inverted microscope. These methods were then used to capture the movement of motile E. coli in Newtonian solutions of Polyethylene Glycol (PEG), and their motility parameters were calculated using DDM. Similarly, preliminary data on the motility of E. coli in aqueous HNP solutions were collected in concentration regimes where the HNP solutions do not exhibit a yield-stress. This thesis establishes a framework for studying E. coli motility in complex fluids, paving the way for examining the effects of yield-stress and viscoelasticity on the motility parameters.