Toluene diisocyanate (TDI)-based polyisocyanate prepolymer was prepared via TDI self-polymerization with different catalysts at various temperatures. The self-polymerization process was followed by gel permeation chromatography to investigate the formation of individual components. The contents of isocyanate group (-NCO) and free TDI monomer were monitored to confirm the transformation of individual components through back-titration and gas chromatography, respectively. A step-growth polymerization mechanism for TDI-based polyisocyanate reaction was proposed. Simultaneously, a mathematical model was established to simulate the contents of all components in the self-polymerization process, and the simulated data were in good agreement with the experimental results. Mass spectrometry, Fourier transform infrared spectroscopy and C-13 nuclear magnetic resonance spectroscopy were employed to identify and confirm the chemical structures of the polyisocyanate prepolymer. The experimental results indicate that several oligomers containing trimer, pentamer, heptamer and higher-molecular-weight oligomers presented simultaneously and interacted with each other. The desired content of each component in TDI-based polyisocyanate prepolymer could be obtained by controlling the reaction conditions.