Comparison of two different tissue decalcification methods for immunohistochemistry

Objective:To investigate the effects of 1% hydrochloric acid and 10% nitric acid on immunohistochemical staining for several antigens.Methods: Fifteen cases of focally calcified soft tumors including three cases of thyroid papillary carcinoma,three of breast invasive ductal carcinoma,three of prostate cancer,three of colon adenocarcincoma and three of normal lymph nodes,were selected in this study,and so were nine cases of bone and bone marrow tissue from biopsy and autopsy.Tissue samples were divided into three groups.One group was each soaked into 10% nitric acid for 24,42 and 48 h,another group was each incubated in 1% hydrochloric acid for 1,2 and 3 h.The other group which was not treated with acid was regarded as control.Then immunohistochemical staining was conducted in these samples for antigens as follows: ER,PR and Ki-67 in breast invasive ductal carcinoma;TTF-1 in thyroid papillary carcinoma;34βE12 and P504s in prostate cancer;AE1/AE3,P504s and Ki-67 in colon adenocarcincoma;CD3,CD20 and Ki-67 in lymph nodes;CD3,CD20,CD68,CD235a,MPO,Ki-67,AE1/AE3 and TTF-1 in bone tissue.Results: After the tissues decalcificated in both acid solutions for some time,the numbers of positive cells decreased and the intensity of immunostaining became weaker,which was proportionate to time.Bone tissue was less influenced than calcified soft tissue.Different antigens showed different sensitivity to the same acid treatment.TTF-1 and Ki-67 were the two most vulnerable antigens among them;while ER,CD3 and CD20 were the least influenced.The effects of decalcification on immunostaining of one antigen were the same in different tissues.Conclusion: In most cases,both hydrochloric acid and nitric acid decalcification can lower the sensitivity of immunohistochemical staining,but have different effects on different antigens.It is better to use 10% nitric acid for decalcification of relatively large bone tissue and to use 1% hydrochloric acid for bone marrow tissues and bone tissues containing much more soft tissue.