The Shigella flexneri outer membrane (OM) protease IcsP (SopA) is a member of the enterobacterial Omptin family of proteases, and cleaves the polarly localised OM protein IcsA that is essential for Shigella virulence. Unlike IcsA, the specific localisation of IcsP on the cell surface is unknown. To determine the distribution of IcsP in the OM, a haemagglutinin (HA) epitope was inserted into the non-essential OM loop 5 of IcsP. Quantum Dot (QD) immunofluorescence (IF) surface labelling to detect IcsP^HA was then undertaken. Quantitative IF microscopic analysis of S. flexneri 2a 2457T treated with and without tunicaymcin to deplete lipopolysaccharide (LPS) O antigen (Oag) showed that IcsP^HA was asymmetrically distributed on the surfaces of both septating and non-septating cells, and that this distribution was masked by LPS Oag in untreated cells. Double QD IF labelling of IcsP^HA and IcsA showed that IcsP^HA preferentially localised to the new pole of non-septating cells, and to the septum of septating cells. The localisation of IcsP^HA in an S. flexneri 2457T strain lacking Oag was also investigated and an equivalent distribution of IcsP^HA was observed. Complementation of the rough LPS strain with rmlD resulted in restored LPS Oag chain expression and loss of IcsP^HA detection, providing further support for LPS Oag masking of surface proteins. Our data presents for the first time the distribution for the Omptin OM protease IcsP, relative to IcsA, and the effect of LPS Oag masking on its detection. The asymmetrical distribution of IcsP suggests that the efficiency of IcsA cleavage is also asymmetrical which supports the role of IcsP in establishing and maintaining IcsA polarity.