Microwave Decalcification Research Articles

Evaluation of structural changes of bone in chronic purulent otitis media

Fragments of bone tissue of the temporal bone, obtained during reconstructive-sanitizing operations in patients with chronic purulent otitis media, were studied by light and electron microscopy. An analysis was made of the degree of structural changes in bone tissue in chronic inflammation at the cellular and tissue levels after a histomorphological study in microwave decalcification. The method of diagnosis reliably allowed to differentiate the diseases characterized by the rarefaction of bone tissue, due to chronic inflammation, and also to determine the processes of transformation of bone tissue. The method of diagnosis reliably allows to differentiate diseases characterized by the rarefaction of bone tissue due to chronic inflammation, as well as to determine the processes of bone tissue transformation, however, the study is time-consuming, long-lasting and expensive.

A quantitative study of comparing the routine decalcification and microwave decalcification methods by using different decalcifying agents

Background: Decalcification is a process of complete removal of calcium salts from mineralized tissues. Microwave decalcification is a novel technique that can accelerate the duration of decalcification compared to the manual method, by increasing the temperature to evaluate the fastest decalcifying agents and methods (routine with Microwave decalcification) in both bone and tooth. Materials and Methods: The study group consisted of 80 hard tissues, 40 teeth and 40 pieces of 1 cm × 1 cm cut bone specimens were decalcified by both routine and microwave method using four decalcifying solutions (five bone pieces and five teeth in each agents for both routine and microwave method). The four decalcifying solutions were 8% nitric acid, 8% formic acid, 8%formal nitric acid, and 14% of ethylenediaminetetraacetic acid (EDTA). Results: Duration of decalcifi cation of bone and tooth by both methods with 8% nitric acid took 10 days for CD and 2 days for MD respectively. Similarly for rest of the acids took more time to decalcify in CD method than MD method. Fourteen percent EDTA gave excellent results followed by 8% formal nitric acid, and 8% formic acid has good results than 8% nitric acid. Conclusion: Microwave processing proved to be an efficient and reliable procedure for the decalcification of bones and tooth. Microwave decalcification yielded quicker and better results compared to the routine methods. Therefore, microwave can serve as a quicker and reliable diagnostic method for a pathologist.

Comparison of Microwave Versus Conventional Decalcification of Teeth Using Three Different Decalcifying Solutions.

Background:In routine histopathology, decalcification of teeth is an essential and important step during tissue processing. The present study was attempted to decalcify teeth using microwave method and to compare it with conventional decalcification method.Aim:To compare microwave decalcification with conventional decalcification method with respect to the speed of decalcification, preservation of tissue structure, and efficacy of staining.Materials and Methods:A total of 72 single-rooted premolars were used for both conventional and microwave decalcification methods. Three different types of decalcifying agents at 5% and 7% were used. Decalcifying agents included nitric acid, formic acid, and trichloroacetic acid, each at 5% and 7%. About 6 teeth were included in each category and were decalcified by both conventional and microwave method. After decalcification, all the teeth were examined macroscopically and microscopically. Cramer's V-test was used to determine the statistical significance.Results:The results have shown that microwave method using 5% and 7% nitric acid were the fastest. Structural details and good staining characteristics were better in teeth decalcified by 5% nitric acid and 5% trichloroacetic acid by both the methods.Conclusions:5% nitric acid by microwave method proved to be the best decalcifying agent as it was fast and gave good structural details and staining characteristics.

A Comparison of Conventional and Microwave Decalcification and Processing of Tooth and Mandibular Bone Specimens.

Any laboratory procedure exposes the technician and the pathologists to the hazardous effects of chemicals. Conventional procedures like decalcification and histo-processing employed in laboratories are labour intense and time consuming thereby delaying the report dispatch. The present study was an attempt to employ a kitchen microwave to hasten the process and facilitate faster and accurate reporting; thus, benefitting the technician, pathologist and the patient. To compare conventional and microwave based decalci-fication, processing and staining of tooth and mandibular bone specimens using 5% nitric acid as decalcifying agent. The sample included formalin fixed 180 tooth specimens (60 incisors, 60 premolars, 60 molars) and 60 mandibular bone specimens (approx 0.5cm each). The hard tissue specimens were subjected to varying combination of conventional and microwave decalcification, processing and staining. The entire procedure was blinded and evaluated by two examiners. Conventional Decalcification (CD), processing and staining produced the utmost quality, though consuming a relatively longer duration. Microwave reduced the total decalcification time by half and retained the diagnostic quality of the specimens. On the contrary the microwave based processing and staining caused significant damage to the tissues rendering sections un-diagnostic. A combination of Microwave Decalcification (MD) followed by Conventional Processing (CP) and staining would be ideal to hasten the overall laboratory time with minimal compromise on tissue quality.

Localization of a Specific Inward Rectifying Potassium Channel Protein in the Human Cochlea

Objectives: Our study aims to determine the immunostaining pattern of Kir4.1 channels in human cochlear tissues. Potassium recycling pathways critical for maintaining cochlear ion homeostasis and the endocochlear potential have been defined largely in animal models. Potassium effluxed from sensory hair cells is taken up by supporting cells and returned to the stria vascularis via two distinct transcellular syncytial networks consisting of epithelial cells and spiral ligament fibrocytes. The inward rectifying potassium channel Kir4.1 appears to be an essential component of this process, as evidenced by murine knockout models lacking an endocochlear potential. Animal models have demonstrated robust Kir4.1 expression in several cell types along the cochlear potassium recycling pathway. However, Kir4.1 immunostaining patterns in the human cochlea remain undefined. Methods: Postmortem human temporal bones were collected through the Hearing Research Program at the Medical University of South Carolina. Temporal bones were fixed within 6 to 11 hours of death and underwent microwave decalcification prior to immunohistochemical staining for Kir4.1. Results: Robust Kir4.1 immunoreactivity was present in strial intermediate cells, outer sulcus root processes and glial cells in Rosenthal’s canal. The distribution of Kir4.1 in the human cochlea was generally similar to that reported in animal models. Conclusions: Our findings suggest that Kir4.1 channels play a critical role in the regulation of potassium recirculation in the human cochlea. Further immunohistochemical analyses are necessary to fully delineate the precise location of Kir4.1 in the complex potassium recycling pathway and elucidate its potential role in lateral wall degeneration and associated hearing loss.

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

Introduction:The microwave oven has been used quite often for tissue processing, but there are very few studies describing its use in decalcification of bone or teeth. In this study we have attempted to decalcify bone and teeth using a microwave oven and compare the process and results with conventional decalcification methods.Aims and objectives:The objectives of the study were to determine and compare routine decalcification with microwave decalcification of bone and teeth using 5% nitric acid, 5% formic acid, and 14% ethylenediaminetetraacetic acid (EDTA) with respect to speed of decalcification, preservation of tissue structure and staining efficacy.Materials and methods:In our study the total sample size used for both routine and microwave decalcification was 30 premolar teeth and 30 pieces of condyles. The three solutions were dilute nitric acid (5%), formic acid (5%), and EDTA (14%). Each set consisting of the same type of premolars and condyles in each of the three decalcifying solutions were used in both manual method and microwave method.Results:The results in the present study confirmed the fact that the microwave method using nitric acid was indeed the fastest decalcifying method needing just about 2 days for condyle and 4 days for premolars, compared with routine decalcification. The results also showed that the overall histological picture was good with EDTA and formic acid irrespective of the methods used. In the routine method, nitric acid gave poor cellular detail when compared with microwave method.Conclusion:With our study we conclude that microwave oven decalcification is faster than routine decalcification irrespective of the decalcifying agents used. The tissue preservation and staining efficacy was good in microwave nitric acid decalcification compared to routine nitric acid decalcification. Both formic acid and EDTA show good tissue preservation and staining efficacy irrespective of the method used.

Effects of Microwave Fixation and Decalcification on Rodent Tissue

The histological use of laboratory microwaves was proposed by Mayers in 1970. Since then, microwaves have become widely used in histology laboratories for fixation, decalcification, and processing; however, protocols must be carefully optimized to ensure the maintenance of optimal cellular morphology and antigenicity. In the current study, we sought to decrease laboratory turnaround time and optimize specimen preservation conditions by using microwave fixation. Mouse and rat tissues were harvested and divided into two groups (A and B). Tissues in group A were fixed for 24 h in 10% neutral buffered formalin (NBF), whereas tissues in group B were fixed for 2 h in 10% NBF before being microwaved at 50°C for varying times. Tissues from the two groups were compared by a pathologist. We found that the optimal microwave formalin-fixation times necessary to preserve morphology were 29 min for mouse tissues and 30 min for rat tissues. These times allowed specimens to be placed on a processor 3 h after tissue collection and processed for embedding the following day. We also determined optimal times for microwave decalcification of mouse and rat sternums and femurs. Decalcification of rat femurs with the combined use of microwave fixation and decalcification saved nearly 2 days of specimen turnaround times. To assess the effect of microwave fixation on antigenicity, rat intestine samples from groups A and B were stained with Ki-67 and caspase 3 antibodies. When compared with tissues that had been bench-fixed for 24 h, microwave-fixed tissues showed no loss of antigenicity. We conclude that rodent fixation and decalcification times can be considerably reduced with the aid of a microwave, thereby decreasing study turnaround times, minimizing costs, and improving efficiency. Microwave fixation would be a valuable investment for any laboratory seeking to save time while maintaining optimal cellular morphology and antigenicity. (The J Histotechnol 32(4):190–192, 2009)

Effects of Microwave Fixation and Decalcification on Rodent Tissue

Effects of Microwave Fixation and Decalcification on Rodent Tissue

Microwave decalcification of human temporal bones.

Morphological and immunohistochemical studies of human temporal bones are challenging as a result of difficulties in obtaining reliably fixed specimens and the lengthy time required for decalcification, typically 4 to 7 months. A novel method of processing human temporal bones using a microwave oven to accelerate decalcification is described. This procedure provides a rapid means of decalcifying temporal bones with good preservation of tissue morphology and antigenicity. Human temporal bone specimens obtained at autopsy (n = 12, from specimens aged 43-91 y) were fixed within 6.5 hours of death by transtympanic perilymphatic perfusion of the inner ear. Decalcification was carried out using ethylenediaminetetra-acetic acid (EDTA) in a microwave oven and required only 3 to 6 weeks. Specimens were then dehydrated, embedded in paraffin, sectioned, and mounted on slides for morphological and immunohistochemical evaluation. Microscopic examination revealed no obvious artifacts attributable to the microwave decalcification process. The quality of morphological preservation was largely dependent on the postmortem fixation interval and adequacy of perilymphatic perfusion. Immunohistochemical analysis demonstrated strong positive staining for the enzyme Na,K-ATPase, an integral membrane protein. This study demonstrates that microwave decalcification provides an efficient and reliable means of processing human temporal bones for histological and histochemical examination. Decalcification time is significantly reduced with no apparent adverse effects on structural preservation or antigenicity.

Immunohistochemistry and microwave decalcification of human temporal bones

Processing of human temporal bones is a long, expensive process and the resulting celloidin sections are difficult to use for immunohistochemistry. We tested the ability of immunohistochemical assays to work in human temporal bones that were decalcified using a microwave oven. Tissue was trimmed to an approximate cube (1.5–2 cm/side) containing only the cochlea and immersed in fresh EDTA with paraformaldehyde every 6 h. This sized block required 190–400 h to decalcify. The decalcified tissue was embedded in paraffin and sectioned. Sections were immunoassayed with anti-cytochrome c oxidase, anti-neurofilament or anti-peripherin. All three antibodies labeled the appropriate structures. This procedure may stimulate advancement in the understanding of human inner ear pathology.

Rapid Decalcification Using Microwaves for in Situ Hybridization in Skeletal Tissues

In situ hybridization histochemistry is the sole tool available for detecting the localization and expression of specific RNA on histological sections under various in vivo conditions. For this paper, we examined the effect of microwave exposure on the time needed for decalcification of skeletal tissues and on the preservation of sensitivity for hybridization signals. Our data show that the use of microwave decalcification reduces the decalcification period while preserving intense hybridization signals for mouse al chain of procollagen type I mRNA in osteogenic cells in bone. The use of microwave treatment to decalcify skeletal tissues may prevent delay in obtaining experimental results or the loss of signals during in situ hybridization.