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ORIGINAL RESEARCH
Year : 2014  |  Volume : 1  |  Issue : 1  |  Page : 3-6

Comparison of conventional and microwave histo-processing of various oral soft tissue specimens


Department of Oral Pathology, M.S. Ramaiah Dental College, Bangalore, Karnataka, India

Date of Web Publication31-Jan-2014

Correspondence Address:
Shankargouda Patil
Department of Oral Pathology, M.S. Ramaiah Dental College, Bangalore, Karnataka
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/2348-3172.126154

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  Abstract 

Background and Aim: While a number of pathologists have reviewed the techniques and results of microwave-facilitated tissue fixation and processing, there has been no record of any previous studies where specific tissues were chosen and compared. Hence, the aim of the present study was to specifically evaluate and compare the diagnostic ability of selective oral soft tissue specimens processed and stained by the conventional and microwave method. Materials and Methods: The study group comprised of 40 formalin-fixed tissue samples from the archives of the Department of Oral Pathology, 10 each of epithelial, muscle, adipose and glandular tissue. Each specimen was cut into two halves with one half processed and stained by the conventional method while the other by the microwave method. With the procedure blinded, four observers evaluated the slides employing Babu et al's criteria pertaining to cellular clarity, cytoplasmic details, nuclear detail and color intensity. The results were statistically analyzed using Chi-square test and kappa. Results: The microwave method yielded better results as compared to the conventional with respect to processing and staining although there was no statistical difference between the two. A drastic reduction in time with the microwave method was observed. Conclusion: The results obtained by microwave method surpassed the conventional method. Hence, it is ideal to adopt the microwave method for any oral soft tissue for quicker and reliable results.

Keywords: Conventional, histo-processing, microwave, oral soft tissues


How to cite this article:
Patil S, Rao RS, Nagaraja A, Kumar SD. Comparison of conventional and microwave histo-processing of various oral soft tissue specimens. J Dent Res Rev 2014;1:3-6

How to cite this URL:
Patil S, Rao RS, Nagaraja A, Kumar SD. Comparison of conventional and microwave histo-processing of various oral soft tissue specimens. J Dent Res Rev [serial online] 2014 [cited 2019 Oct 17];1:3-6. Available from: http://www.jdrr.org/text.asp?2014/1/1/3/126154


  Introduction Top


In order to preserve the structure of any tissue and impregnate them with a suitable media, they have to be adequately fixed and processed, so that thin sections can be made for staining and microscopic evaluation. However, the legacy of traditional way of histo-processing is followed since ages. Although these procedures are elaborate and time consuming, the conventional method is still in vogue. [1]

Microwaves were invented by Percy Spencer in 1945 and soon became an integral part of our daily lives. The potential application of microwave in histotechnology was first recognized in 1970 by Mayers. [2] The advantage microwave offers in histo-processing is that it helps in generating heat from within, uniformly warming the object thereby permitting chemicals to diffuse faster and reduces the overall time required for processing, thus, eliminating the exposure to noxious agents like xylene. [3]

The purpose of the present study was to evaluate and compare various tissues processed and stained by conventional and microwave methods, since there are very few studies pertaining to the same.


  Materials and Methods Top


The materials used were,

  • Microwave oven (Samsung Model no: CE104VD, Input - 1250 W, Output - 900 W)
  • Microwave glass jar - 5 jars 200 ml each
  • Ethyl alcohol
  • Isopropyl alcohol
  • Xylene
  • Paraffin wax
  • Harris hematoxylin, 0.5% HCl.


Sample selection

  • 40 tissue samples from the Department of Oral pathology M.S Ramaiah Dental College and Hospital, Bangalore (samples listed in Flowchart 1 [Additional file 1])
  • Scalpel biopsies (1 × 1 cm up to 3 cm in size, 5-8 mm thickness) fixed in 10% buffered formalin
  • Tissues excluded - Grossly lacerated tissues, hard tissues and surgical procedures other than scalpel biopsies.


One half of each specimen was processed and stained by conventional method (as per the department protocol - [Table 1]) and the other half by the microwave method (as per Babu [2] et al. - [Table 2]). The microwave temperature was standardized at 100 W.
Table 1: Conventional and microwave processing techniques


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Table 2: Conventional and microwave staining techniques


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The entire procedure was blinded and evaluated by four observers using criteria by Babu et al. [2] [Table 3] and analyzed using Chi-square test and kappa statistics.
Table 3: Criteria for evaluation of quality of slides


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  Results Top


Epithelial, muscle, adipose and glandular tissues from [Table 4], [Figure 1] and [Figure 2] were better appreciated when processed and stained by microwave processing and staining, as compared to the conventional processing and staining method. The P value was 0.066, depicting that there was no statistical difference between the two methods and kappa statistics showed high agreement between the observers.
Figure 1: Microwave (capital letters) verses conventional method (small letters). (A, a) The Cellular clarity, nuclear details, cytoplasmic details and color intensity noted in the epithelium, (B, b) Muscle, (C, c) Adipose, (D, d) Mucous gland, (E, e) Serous gland

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Figure 2: Representation of influence of microwave and conventional processing and staining on various tissues

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Figure 3: Representation of time taken for microwave and conventional processing and staining

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Table 4: Mean scores obtained for microwave and conventional processing and staining on various tissues


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The time taken as per from [Figure 3] for microwave processing and staining was considerably less, when compared to the conventional protocol.


  Discussion Top


The introduction of microwave has brought about reassessment of traditional concepts. [2] The mechanism of microwave heating depends on oscillating or exciting polar or charged molecules. Alternating electromagnetic fields are produced which cause polar molecules of proteins to rotate through 180° at 2.45 billion cycles per second. These result in generation of instantaneous heat that is proportional to the energy flux and continues until the radiation ceases. The microwaves stimulate the polar molecules causing collision with the adjacent molecules which causes part of the rotational energy to be transferred through them producing heat. This effect occurs simultaneously throughout the whole material being microwaved. [3]

The usage of microwave provides shorter processing time, lesser degree of denaturation of nucleic acids and removal of noxious chemicals like xylene. [4],[5] Also, domestic microwaves are readily available, affordable and have provided appreciable results in previous studies. [3] Hence, the same was opted for the present study.

The microwave was operated at a power of 100-300 W for histoprocessing and staining in order to prevent tissue damage. This is in conjunction with the studies of Rajuet et al. [3] and Babu et al. [2] Also, microwave glass wares were preferred as metallic utensils ignite sparks, favoring Babu et al.'s [2] study.

Six tissues/load with a uniform thickness of 5-8 mm were processed which favored adequate and proper diffusion, although the maximum tissue load permissible is up to 25 samples/load. [6] This was in accordance with Prasad et al.'s [1] study which reflects that diffusion is dependent on the thickness and not on the length and breadth of the specimen.

The steps involved in conventional processing are elaborate while the microwave method are much simpler and employ isopropyl alcohol alone with dual effect of dehydrant and clearing as the residual alcohol gets evaporated by the microwave energy during impregnation, thereby eliminating the need for a separate clearing procedure. This in turn cuts the cost, materials and the hazardous effects of xylene. This is in accordance with the study of Pritam and Raju et al. [2],[4]

We observed the effect of microwave processing and staining on various types of oral tissues, and to the best of our knowledge there are very few studies with respect to this.

The epithelial tissue [Figure 1]a processed and stained by the microwave method showed better quality of cellular and nuclear details with prominent intercellular bridges. These findings were in agreement with Panja and Boon et al.'s study. [4],[7]

Microwave processed and stained salivary gland tissue [Figure 1]d containing mucous and serous cells [Figure 1]e have shown better quality of cytoplasm with its contents and crisp nuclear staining when compared to the conventional method. Similar results were appreciated in the Panja et al.'s study. [4]

Adipose tissue [Figure 1]c, muscle tissue [Figure 1]b also showed good preservation of cellular, nuclear details in tissues processed and stained by the microwave method and to the best of our knowledge we have not found any literature relating to these tissues. Hence, we can confidently use the microwave processing and staining method for the above-mentioned tissues without any speculation, as the results obtained have excelled over the other.

Also, most significantly we noted that microwave was less time consuming (7 h 31 min 20 sec) when compared to the conventional protocol (2 h 16 min 45 sec) [Figure 3]. This would permit diagnosis on the same day and facilitate management on a 1-day basis. This was in agreement with the studies conducted by Ralph et al., Morales et al. and Babu et al. [2],[4],[5],[8]


  Conclusion Top


The merits of microwave histo-processing have surpassed the routine conventional protocol in many ways like: Being less labor intensive, less toxic and facilitating rapid diagnosis. Despite the many advantages of microwave, we continue to adhere to routine formalin fixation and conventional processing as the "gold standard" for histological examination against new technologies. Though greater caution is to be exercised while the microwave is handled, using microwave technology in the laboratory on a regular basis is the need of the hour.

 
  References Top

1.Kango PG, Deshmukh RS. Microwave processing: A boon for oral pathologists. J Oral Maxillofac Pathol 2011;15:6-13.  Back to cited text no. 1
[PUBMED]  Medknow Journal  
2.Babu TM, Malathi N, Magesh KT. A comparative study on microwave and routine tissue processing. Indian J Dent Res 2011;22:51-5.  Back to cited text no. 2
    
3.Shashidara R, Sridhara SU. Kitchen microwave-assisted accelerated method for fixation and processing of oral mucosal biopsies: A pilot study. World J Dent 2011;2:17-21.  Back to cited text no. 3
    
4.Panja P, Sriram G, Saraswathi TR, Sivapathasundharam B. Comparison of three different methods of tissue processing. J Oral Maxillofac Pathol 2007;11:15-7.  Back to cited text no. 4
  Medknow Journal  
5.Rohr LR, Layfield LJ, Wallin D, Hardy D. Comparison of routine and rapid microwave tissue processing in a surgical pathology laboratory quality of histologic sections and advantages of microwave processing. Am J Clin Pathol 2001;115:703-8.  Back to cited text no. 5
[PUBMED]    
6.Devi RB, Subhashree AR, Parameaswari PJ, Parijatham BO. Domestic microwave tissue processing. J Clin Diagn Res 2013;7:835-9.  Back to cited text no. 6
    
7.Kok LP, Boon ME, Suurmeijer AJ. Major improvement inmicroscopic-image quality of cryostat sections: Combining freezing and microwave-stimulated fixation. Am J Clin Pathol 1987;88:620-3.  Back to cited text no. 7
[PUBMED]    
8.Morales AR, Nassiri M, Kanhoush R, Vincek V, Nadji M. Experience with an automated microwave-assisted rapid tissue processing method validation of histologic quality and impact on the timeliness of diagnostic surgical pathology. Am J Clin Pathol 2004;121:528-36.  Back to cited text no. 8
[PUBMED]    


    Figures

  [Figure 1], [Figure 2], [Figure 3]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]



 

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Abstract
Introduction
Materials and Me...
Results
Discussion
Conclusion
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