Abstract:

Background: Bones are hard tissues forming the body skeleton, composed of 60% inorganic component (hydroxyapatite), 10% water and 30% organic component. In routine histopathology, the study of fibrillar, cellular and sub cellular structures of mineralized tissues like bones is only possible after removal of the calcium apatite of these hard tissues by the process of decalcification using different decalcifying agents such as acids or chelating agents. In our setting there is no study about comparing the efficacy of 10% formic acid and cocktail mixture of nitric acid and EDTA. The aim of the study was changing 10% formic acid the routine decalcifying agent to cocktail mixture.

Objectives: The study evaluated the efficacy of two different decalcifying agents by identifying fastest decalcifying agents, effects of agents on integrity of the bone tissues, and staining characteristics of the bones decalcified in the two different decalcifying agents.

Material and methodology: Prospective descriptive study was conducted on all bone tissues in Histopathology and Morbid anatomy Laboratory at BMC from June to July 2024. Total of 56 bone cases were studied which are skulls, ribs, clavicles, scapulars and teeth. For each case, that was obtained at histopathology laboratory was well fixed, then decalcified (using two decalcifying solutions were prepared), processed, embedded, sectioned, stained and observed microscopically according to BMC-Histopathology and morbid anatomy laboratory standard guidelines. The scores were given on five parameters: speed of decalcification, background, overall staining pattern, cell morphology and nuclear staining.

Results: A total number of 56 bone tissue samples were enrolled in this study, cocktail of Nitric acid and EDTA was the fastest agent which took 9.8 ± 6 days while 10% formic acid was slowest agent 15.2 ±5 days for complete decalcification, 10% formic acid shown best results compared to cocktail mixture on other parameters which are overall staining, background, cell morphology and nuclear character.

Conclusion: Histological sections impressions depend on a variety of factors which includes fixation, tissue processing, sectioning and staining timings. The final results led to the proposition that 10% formic acid was indeed the best decalcifying agent and should be used if time factor is not essential. In case of time become relevant for example if surgeon depend on pathologist output for patient management, cocktail of 10% nitric acid and 1% EDTA can be used due to its fastest action and good staining features.