Microwave-induced fast decalcification of rat bone for electron microscopic analysis: an ultrastructural and cytochemical study.
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.
Pitol
Dimitrius Leonardo
DL
Department of Morphology, Stomatology and Physiology, School of Dentistry of Ribeirão Preto, University of São Paulo, Ribeirão Preto, SP, Brazil. dimipitol@yahoo.com.br
Caetano
Flavio Henrique
FH
Lunardi
Laurelúcia Orive
LO
D000818:Animals; D001857:Bone Matrix / Q000528:radiation effects / Q000648:ultrastructure; D001842:Bone and Bones / Q000528:radiation effects / Q000648:ultrastructure*; D002118:Calcium; D002614:Chelating Agents; D003080:Cold Temperature; D003094:Collagen / Q000528:radiation effects / Q000648:ultrastructure; D003461:Crystallography; D003648:Decalcification Technique; D004492:Edetic Acid; D005404:Fixatives; D005976:Glutaral; D008437:Maxilla / Q000528:radiation effects / Q000648:ultrastructure; D046529:Microscopy, Electron, Transmission; D008872:Microwaves; D015388:Organelles / Q000528:radiation effects / Q000648:ultrastructure; D010010:Osteoclasts / Q000528:radiation effects / Q000648:ultrastructure; D010011:Osteocytes / Q000528:radiation effects / Q000648:ultrastructure; D051381:Rats; D017208:Rats, Wistar; D012972:Sodium Hydroxide; D013048:Specimen Handling / Q000379:methods; D013997:Time Factors
D002614:Chelating Agents; D005404:Fixatives; D012972:Sodium Hydroxide; D003094:Collagen; D004492:Edetic Acid; D002118:Calcium; D005976:Glutaral