Microwave-induced fast decalcification of rat bone for electron microscopic analysis: an ultrastructural and cytochemical study

Abstract

Bone decalcification is a time-consuming process. It takes weeks and preservation of the tissue structure depends on the quality and velocity of the demineralization process. In the present study, a decalcification methodology was adapted using microwaving to accelerate the decalcification of rat bone for electron microscopic analysis. The ultrastructure of the bone decalcified by microwave energy was observed. Wistar rats were perfused with paraformaldehyde and maxillary segments were removed and fixed in glutaraldehyde. Half of specimens were decalcified by conventional treatment with immersion in Warshawsky solution at 4 degrees C during 45 days, and the other half of specimens were placed into the beaker with 20 mL of the Warshawsky solution in ice bath and thereafter submitted to irradiation in a domestic microwave oven (700 maximum power) during 20 s/350 W/+/-37 degrees C. In the first day, the specimens were irradiated 9 times and stored at 40 degrees C overnight. In the second day, the specimens were irradiated 20 times changing the solution and the ice after each bath. After decalcification, some specimens were postfixed in osmium tetroxide and others in osmium tetroxide and potassium pyroantimonate. The specimens were observed under transmission electron microscopy. The results showed an increase in the decalcification rate in the specimens activated by microwaving and a reduction of total experiment time from 45 days in the conventional method to 48 hours in the microwave-aided method.

Comparison of microwave decalcification with conventional decalcification method by using different decalcifying agents

“…A new method using microwave oven was seen to accelerate the decalcification. 6 The choice of decalcifying agent and method is largely dictated by the urgency of the procedure. 7 The potential application of microwave energy in histotechnology was first recognized by Mayers (1970).…”

“…Another study has shown a reduction of decalcification time from 45 days to 2 days when done by microwave method. 6 Others opinion that the bone decalcification is accelerated by about 10 times using microwave method. 9 The results of the present study were consistent with the previous studies, concluding that the microwave-assisted method is faster than the conventional method.…”

Comparison of routine decalcification methods with microwave decalcification of bone and teeth

“…A new method using microwave oven was seen to accelerate the decalcification. [6] The choice of decalcifying agent and method is largely dictated by the urgency of the procedure. [7]…”

“…This energy induces a dielectric field leading to a rapid oscillation of dipolar molecules at about 180°C, generating heat that is rapidly distributed homogeneously within the tissue. [6]…”

Excesso de tiroxina materna associado ao hipertireoidismo pós-natal reduz o crescimento ósseo e o perfil proliferativo e angiogênico das cartilagens de crescimento de ratos

“…Posteriormente, os ossos foram colocados na segunda solução de EDTA a 10%, em água destilada, até completa descalcificação (Pitol et al, 2007). Em seguida, os ossos foram processados pela técnica rotineira de inclusão em parafina e submetidos à microtomia para obtenção de secções histológicas de 4μm, coradas por HE.…”