Single-walled carbon nanotubes (SWNTs) are suitable for many applications in molecular imaging because of their intrinsic near-infrared (NIR) photoluminescence. Nevertheless, several important issues, such as the SWNTs low biocompatibility, still need to be overcome for the effective use of carbon nanotubes in molecular imaging. In this study, we employ a two-step oxidation procedure with the aims of (1) removing the toxic metal impurities, (2) decreasing the lengths of the tubes, and (3) introducing functional oxygenated groups on the SWNT surface to enable the future development of a multifunctional SWNT-based nanoplatform. Since the presence of defect sites on the SWNT surface is known to quench the optical properties of carbon nanotubes, we have established what effect these two chemical treatments have on the NIR photoluminescence (PL) of the tubes. Our results show that purified and oxidized SWNT samples still display structured emission features in the NIR when dispersed in surfactant aqueous solution, with a residual emission efficiency of 30%. Two-dimensional (2D) fluorescence lifetime imaging (FLIM) of aggregates of purified and oxidized SWNTs in dimethylformamide (DMF) showed an average lifetime of 1.3 and 1.2 ns, respectively. These results demonstrate the relevance of SWNT purification and oxidation in the development of novel NIR probes with reduced toxicological impact on human health and with potential for further functionalization with multiple bioactive species.