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Objectives: This study aims to clarify the cause of background coloration in the epithelia between each dilated intra papillary capillary loop in esophago-pharyngeal squamous cell carcinoma. Design: This is a single center retrospective study including 124 patients with 160 lesions who underwent esophagogastroduodenoscopy in Nagasaki University Hospital from September 2007 to March 2012; a detailed comparison between endoscopic images and pathology was performed. Immunohistological assessment using anti-human hemoglobin antibody (anti-Hb Ab) was performed to verify the presence of hemoglobin (Hb) component in the cancer cells. Real-time polymerase chain reaction (RT-PCR) and in situ hybridization (ISH) on Hb-β mRNA were performed to assess the production of Hb component within the cancer cells. Results: A strong positivity for anti-Hb Ab was observed in the squamous cell carcinoma area, whereas non-cancerous mucosa showed no immunopositivity for Hb. The concordance rate between anti-Hb Ab immunoreactivity and the presence of BC was as high as 80.9%. The amount of Hb-β mRNA expression was three times higher in cancer tissues compared with the surrounding non-cancerous mucosa. ISH images showed that the expression exclusively occurred in cancer cells, indicating that Hb is probably produced within cancer cells. Conclusions: The background coloration observed is partly due to an extravascular component of Hb. RT-PCR and ISH analyses indicate that Hb is produced within cancer cells. [ABSTRACT FROM AUTHOR]