Immunohistochemistry (IHC) is an indispensable method for the diagnosis of malignancy when alterations in tissue morphology remain deceptively subtle. For more than forty years, antibodies have demonstrated efficacy in chromogenic IHC. However, antibodies continue to harbour issues of batch-to-batch variability and cross-reactivity that are of detriment to the accuracy and precision of clinical histopathology. In contrast, aptamers are a relatively recent class of oligonucleotide-based, bio-recognition molecules, also known as ‘chemical antibodies.’ Aptamers are sequence-defined and synthesised entirely in vitro – virtually eliminating concerns of batch-to-batch variability, whilst providing ease of functionalisation. In addition, aptamers are around 10-20 times smaller than antibodies, which may prove beneficial in detecting cells which co-express multiple surface markers of tumourigenicity. Currently, as a result of the large size of the antibody-chromogen complex, and the resulting potential for steric hindrance, no chromogenic, immunohistochemical double -staining systems are recommended for the detection of bio-markers which are co-localised within the same cellular compartment. We sought to utilise ‘aptahistochemistry’ to overcome this limitation. Having previously generated aptamers against the cancer stem cell markers EpCAM and CD133, we tested their sensitivity in a chromogenic, sequential double-staining technique in formalin-fixed paraffin-embedded colorectal carcinoma xenografts. We were able to demonstrate the detection of cells which co-express both markers at the cell surface. Given that the majority of cells in the bulk tumour show a moderate expression of each of these markers, parallel detection of both EpCAM and CD133 enhances the likelihood that highly co-expressing cells are cancer stem cells – which represent an attractive therapeutic target. The ability to sensitively detect co-localised bio-markers, without concerns over batch-to-batch variation, points to a promising future for aptamers in the fields of clinical histopathology and personalised oncology. However, attention to subtle differences in histochemical methodology are required for these non-protein reagents - particularly in regard to the use of anionic competition to both enhance sensitivity and block non-specific ionic interactions of the negatively charged aptamers.