Immunohistochemistry (IHC) and immunofluorescence (IF) are two most often used tools in the researches of tissue morphology and cell biology. In order to demonstrate which is more sensitive in the detection in paraffin embedded tissues (PET), we compared the detection rate of these two methods with human pancreatic tissue microarrays (TMA) by using three anti-human insulin antibodies. The results showed that the detection rates are mostly depending on the primary antibody (Ab) chosen. For some primary Abs, there were no significant differences between IHC and IF in PET tissues, such as primary Ab No. 1 (85.7%, 18/21 vs 76.2%, 16/21) and Ab No. 3 (95.2%, 20/21 vs 95.2, 20/21). In contrast, for other primary Abs (No. 2), the detection rate of IF is much lower than that of IHC in PET tissues (38.1%, 8/21 vs 90.5%, 19/21). Interestingly, the detection rate of IF was increased after the same sections were re-stained with the procedure of IHC. In addition, our results also showed that IHC method can be executable after IF method was finished in the same PET tissues, while it is hardly to execute IF after the DAB staining of IHC method was completed. We conclude that whether IF or IHC method can be applicable in the PET tissues largely depend on the primary Abs you have chosen. Also, we found that IHC method can still be executable after IF method was finished in the same PET tissues.