Background Although macular amyloidosis is a relatively rare disease, it is a common cutaneous disease in Asia and the Middle East. On hematoxylin and eosin (H&E) stained slides, early lesions could easily be missed without the use of special stains and/or immunohistochemistry. Methods We enrolled 42 patients with the clinical impression of macular amyloidosis who had undergone two 4-mm punch biopsies from 2015 to 2016 at a dermatology clinic affiliated to Shiraz University. Besides, 14 cases with a clinical diagnosis other than macular amyloidosis were selected as the negative control group. Congo red, crystal violet, and immunohistochemical (IHC) staining of CK5 and high molecular weight keratin (HMWK) were performed for each specimen. Results H&E slides showed globular depositions in 15 (35.7%) out of 42 patients. None of the patients showed apple-green birefringence with Congo red stain. Evaluation of crystal violet stained sections revealed purplish violet amyloid deposits in 15 (35.7%) patients. IHC study showed expression of CK5 in 52.4% and HMWK in 50% of the patients, which was not a significant difference (p = 0.715). The findings of both IHC markers had a significant difference with H&E stains (p = 0.039) and crystal violet (p = 0.008). Additionally, we found that two punch biopsies from two sites in the involved area did not have a significant preference over one punch biopsy. All of the cases in the control group were negative for amyloid deposition in H&E, special stains, and IHC stained slides as expected. Conclusions IHC evaluation using CK5 and HMWK might be a useful tool for diagnosing macular amyloidosis.