Use of tumor-targeted fluorescence dyes to help surgeons identify otherwise undetected tumor nodules, decrease the incidence of cancer-positive margins, and facilitate localization of malignant lymph nodes has demonstrated considerable promise for improving cancer debulking surgery. Unfortunately, the repertoire of available tumor-targeted fluorescent dyes does not permit identification of all cancer types, raising the need to develop additional tumor-specific fluorescent dyes to ensure localization of all malignant lesions during cancer surgeries. By comparing the mRNA levels of the hypoxia-induced plasma membrane protein carbonic anhydrase IX (CA IX) in 13 major human cancers with the same mRNA levels in corresponding normal tissues, we document that CA IX constitutes a nearly universal marker for the design of tumor-targeted fluorescent dyes. Motivated by this expression profile, we synthesize two new CA IX-targeted near-infrared (NIR) fluorescent imaging agents and characterize their physical and biological properties both in vitro and in vivo. We report that conjugation of either acetazolamide or 6-aminosaccharin (i.e., two CA-IX-specific ligands) to the NIR fluorescent dye, S0456, via an extended phenolic spacer creates a brightly fluorescent dye that binds CA IX with high affinity and allows rapid visualization of hypoxic regions of solid tumors at depths >1 cm beneath a tissue surface. Taken together, these data suggest that a CA IX-targeted NIR dye can constitute a useful addition to a cocktail of tumor-targeted NIR dyes designed to image all human cancers.