The filamentous cyanobacterium Nostoc sp. strain PCC 7120 is capable of fixing atmospheric nitrogen. The labile nature of the core process requires the terminal differentiation of vegetative cells to form heterocysts, specialized cells with altered cellular and metabolic infrastructure to mediate the N2-fixing process. We present an investigation targeting the cellular proteomic expression of the heterocysts compared to vegetative cells of a population cultured under N2-fixing conditions. New 8-plex iTRAQ reagents were used on enriched replicate heterocyst and vegetative cells, and replicate N2-fixing and non-N2-fixing filaments to achieve accurate measurements. With this approach, we successfully identified 506 proteins, where 402 had confident quantifications. Observations provided by purified heterocyst analysis enabled the elucidation of the dominant metabolic processes between the respective cell types, while emphasis on the filaments enabled an overall comparison. The level of analysis provided by this investigation presents various tools and knowledge that are important for future development of cyanobacterial biohydrogen production.