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A comparative study on microwave and routine tissue processing
Abstract
The present study was aimed at assessing the rapid microwave-assisted tissue processing and staining to determine if it can replace standard formalin-fixed and paraffin-embedded processing and staining technique.
The study group consisted of 15 oral mucosal biopsies. The specimens were fixed in 10% formalin for 24 hours to ensure adequate fixation and their gross features were recorded, photographed, and then the specimens were cut into equal halves to be processed by both conventional and microwave histoprocessing methods and then subsequently stained with H and E by microwave and conventional methods. The stained slides in each group processed by both microwave and routine methods were randomly numbered for a blind study and circulated among six observers. All the observers were asked to grade each parameter into Excellent/Good/Average/Poor in a data sheet comprising a total of six parameters. These gradings were given a numerical value of 4, 3, 2 and 1, respectively. The parameters included in the data sheet were cellular clarity, cytoplasmic details, nuclear details, color intensity, interface of epithelium and connective tissue. Wilcoxon-matched pairs signed rank test (non-parametric) was used to calculate the test of significance (P value).
The total processing time involved in microwave was 42 minutes and 270 minutes for the conventional method. H and E staining in microwave took 33 minutes and 40 minutes for conventional method.
The individual scores by different observers regarding the various parameters included in the study were statistically insignificant, the overall quality of microwave-processed and microwave-stained slides appeared slightly better than conventionally processed and stained slides.
... This will help in rapid patient diagnosis and management. [5] The kitchen microwave oven improvement will reduce not only the turnaround time, but also the costs associated with diagnosis and less chances for patient dissatisfaction, which by default could lead to hastening the initiation of therapy, especially in neoplastic diseases. ...
... On literature review, there are many studies which have analyzed the microwave processing of tissue [3][4][5][6] and only few studies which have evaluated the microwave staining technique. [6,7] In all of these studies, the sample size was small. ...
... We did not find significant difference in nuclear as well as cellular characters by the microwave method in processing [ Table 3]. Our findings are consistent with the studies done by Babu et al., [5] Shashidara et al., [13] Mathai et al., [14] Kumar et al., [15] and Choji et al. [16] The physicochemical basis of tissue processing lies in the diffusion of reagents into the tissue to be processed. Microwaves, which are formed due to nonionizing radiation, produce alternating electromagnetic fields that result in the rotation of polar molecules such as water and the polar side chains of proteins through 180° at the rate of 2.45 billion cycles/s. ...
Background
Histopathological diagnosis of a specimen requires good histoprocessing. It ultimately depends on sample preparation and staining, which is time consuming by routine tissue processing technique. The objective of the present study was to compare the reliability of kitchen microwave for tissue processing and staining with the standard conventional processing and staining in different tissue samples.
Materials and Methods
The study sample included 204 specimens from different parts of the body. Three set of tissues were processed and three set of slides from each sample were stained by routine as well as microwave method and the quality of microscopic features was compared.
Results
No significant difference was found in nuclear and cellular characters in case of staining by both methods. While, in case of processing, mean scoring for nuclear characters for routinely processed and microwave-stained (RM) slides was 6.95 ± 0.09 (confidence interval [CI] 6.83–0.09), for microwave-processed and microwave-stained (MM) slides, it was 6.82 ± 0.07 (CI 6.74–6.90) by observer 1 ( P = 0.068). The mean score of cellular character of RM slides was 7.85 ± 0.06 (CI 7.76) and for MM slides, it was 7.65 ± 0.17 (CI 7.47–7.88) by observer 2 ( P = 0.076). The microwave method was better for both processing ( P = 0.001) and staining ( P = 0.003) in case of small biopsies ( P = 0.057). The microwave method was as good as routine method for processing of aorta and staining of breast and kidney tissues ( P = 0.23, 0.32, and 0.38, respectively). The time taken by the microwave method is considerably less for both processing and staining.
Conclusion
The microwave method requires less time when compared with the routine method, thus reducing the overall turnaround time which ultimately helps in rapid same-day diagnosis and patient management. Microwave can be preferred method for smaller biopsies when compared with large biopsies. Lack of automation and requirement of more manpower remain the major disadvantages for use of microwave method in large turnover histopathology laboratories.
... Similarly when Mathai et al. [8] compared the nuclear size shape and quality in conventional and microwave processed tissues did not find any significant difference. Whereas Babu et al. [9] and Kango and Deshmukh [10] found microwave processing to be better than conventional processing when nuclear detail is concerned. ...
... When the cytoplasmic details in the epithelium, connective tissue, or the overall tissue were evaluated, none of the three observers found any statistical difference between the conventional and microwave processing which was in accordance to various studies. [5][6][7][8][9][10] Babu et al. [9] noticed more shrinkage in microwave processed tissues whereas Panja et al. [4] reported less shrinkage.. Kango and Deshmukh [10] reported uneven staining as an artifact in conventional processing. However, in our study, both the processes of tissue processing that is conventional processing and microwave processing did not show any difference based on the observations given by the observers in the occurrence of artifacts. ...
... When the cytoplasmic details in the epithelium, connective tissue, or the overall tissue were evaluated, none of the three observers found any statistical difference between the conventional and microwave processing which was in accordance to various studies. [5][6][7][8][9][10] Babu et al. [9] noticed more shrinkage in microwave processed tissues whereas Panja et al. [4] reported less shrinkage.. Kango and Deshmukh [10] reported uneven staining as an artifact in conventional processing. However, in our study, both the processes of tissue processing that is conventional processing and microwave processing did not show any difference based on the observations given by the observers in the occurrence of artifacts. ...
Introduction:
Conventional tissue processing takes a lot of time, but microwave processing method though rapid, its quality of tissue texture and staining properties is still questionable. Therefore, we conducted this study to find out the reliability of histological sections in the microwave method.
Aim and objectives:
The aim is to assess whether microwave tissue processing, which is a rapid technique, is comparable to conventional tissue processing in terms of quality and reliability of histological sections.
Materials and methods:
A total of 80 formalin-fixed tissue samples were taken and were divided into two pieces. One subjected to conventional tissue processing and the other subjected to microwave tissue processing. The slides were coded and evaluated by three independent observers using various parameters. All the data were subjected to Chi-square test.
Results:
Both the process did not differ in their efficiency in overall nuclear details, overall cytoplasmic detail, occurrence of artifact and tissue architecture, nuclear staining, cytoplasmic staining, and red blood cell staining. Although the other observers did not find any statistical significance, according to observer 3, conventional processing proved to have marginally significantly better epithelial connective tissue interface than microwave processing (P = 0.047).
Conclusion:
The microwave method of tissue processing is an economical procedure which reduces the obnoxious chemicals as well as the turnaround time.
... Vol. 11, Issue 2 ( April-June 2017) formalin; after that, tissue was wrapped in a fi lter paper and dehydrated in 95% isopropyl alcohol, 100% isopropyl alcohol (Qualikems, Vadodara, Gujarat, India) for 20 min on a stir plate. The dehydrated tissue was cleared in xylene (Qualikems, Vadodara, Gujarat, India) for 20 min on a stir plate. ...
... 6,10 Vol. 11, Issue 2 ( April-June 2017) The applications of microwaves are extensive which includes tissue fi xation, stabilization of large specimens, tissue processing for light and electron microscopy, and histochemical and immunohistochemical staining. Microwave tissue processing technique was introduced by Boon and Kok in 1985, but the potential application of microwave energy was fi rst recognized by Mayers in 1970, who successfully fi xed tissue with a microwave generator. ...
... Vol. 11, Issue 2 ( April-June 2017) and Deshmukh. 25 Since our criterion was adopted from the above-mentioned study, we also placed focal condensation of stroma as indistinct. ...
Objectives
Tissue processing for years is carried out by the conventional method, which is a time-consuming technique resulting in 1-day delay in diagnosis. However, in this area of modernization and managed care, rapid diagnosis is increasingly desirable to fulfill the needs of clinicians. The objective of the present study was to compare and determine the positive impact on turnaround times of different tissue processing methods by comparing the color intensity, cytoplasmic details, and nuclear details of the tissues processed by three methods.
Methods
A total of sixty biopsied tissues were grossed and cut into three equal parts. One part was processed by conventional method, second by rapid manual, and third by microwave-assisted method. The slides obtained after processing were circulated among four observers for evaluation. Sections processed by the three techniques were subjected to statistical analysis by Kruskal–Wallis test. Cronbach’s alpha reliability test was applied to assess the reliability among observers. One-way analysis of variance (ANOVA) was used for comparing mean shrinkage before and after processing.
Results
All observers were assumed to be reliable as the Cronbach’s reliability test was statistically significant. The results were statistically non-significant as observed by Kruskal–Wallis test. One-way ANOVA revealed a significant value on comparison of the tissue shrinkage processed by the three techniques. The histological evaluation of the tissues revealed that the nuclear-cytoplasmic contrast was good in tissues processed by microwave, followed by conventional and rapid manual processing techniques. The color intensity of the tissues processed by microwave was crisper, and there was a good contrast between the hematoxylin and eosin-stained areas as compared to manual methods.
Conclusion
The overall quality of tissues from all the three methods was similar. It was not feasible to distinguish between the three techniques by observing the tissue sections. Microwave-assisted tissue processing has reduced the time from sample reception to diagnosis, thus enabling the same-day processing and diagnosis.
... 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. ...
... 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. ...
... 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. ...
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
... Rapid processing and staining of histological specimen is of paramount importance for early diagnosis and management on a 1 day basis. 4 Diffusion is the key factor in histoprocessing permitting chemicals to permeate into a tissue faster thereby reducing the time. Microwave oven generates heat from within (internal heating) and warms the object uniformly and hence hastens tissue processing. ...
... Hence, the practice of microwave assisted tissue processing, has brought about a revolution and has become a boon to pathologists. 4 Though, there are many studies done on microwave assisted tissue processing, there are only a handful of studies based on microwave staining, hence this study was undertaken to evaluate and compare microwave tissue processing and staining with the conventional method. ...
... The procedure for conventional processing and staining (Tables 1 and 2) was according to our department protocol while microwave processing, and staining (see Tables 1 and 2) was as per Mahesh babu et al (2011). 4 The microwave temperature was standardized at 100 W. ...
Background and aim:
Pathologists are under constant pressure for instant and reliable diagnosis. The manual procedures employed in private laboratories and institutional setup for histoprocessing and staining are laborious and intense. Thus, this study aims to evaluate and compare the microwave tissue processing and staining with the conventional methods which are in vogue.
Materials and methods:
Of the formalin fixed tissue biopsies received by our department, 30 specimens were randomly picked and subjected to grossing. Each specimen was cut into equal halves, each half was processed and stained by conventional method while the other by the microwave method. The entire procedure was blinded and evaluated by four observers based on the criteria of Mahesh Babu et al (2011): Cellular clarity, cyto-plasmic details, nuclear detail and color intensity. The results were statistically analyzed using Chi square test and kappa.
Results:
The overall time employed for microwave processing was 2 hours and for conventional methods it was 7 hours, while H and E staining by microwave process took 16 minutes and 45 seconds and it took 31 minutes and 20 seconds by the conventional process. The diagnostic ability of microwave method yielded promising results and was less time consuming.
Conclusion:
Microwave processing and staining yielded quicker and better results compared to the routine methods. Therefore, Microwave can serve as a quicker and a reliable diagnostic method for a pathologist.
... In a study done by Babu et al., cellular clarity, cytoplasmic details, nuclear details and color intensity were slightly better in microwave method than in routine method. [10] Similar studies by various authors showed no significant differences in the histologic quality when they compared these two protocols. [9,[11][12][13] From the perspective of the end product, microwave irradiation substantially shortens the time from specimen reception to diagnosis. ...
... This is in agreement with Rohr et al. [3] where histological slides were produced in 2-3 h using microwave irradiation. In a study done by Babu et al., [10] the turn around time for microwave tissue fixation and processing and staining was 75 min. In a study done by Henwood A, [14] the tat for microwave tissue fixation and processing was 80 min. ...
The article focuses on whether microwave tissue processing and fixation on tissues of different thickness can replace regular tissue processing and fixation
... In a study done by Babu et al., cellular clarity, cytoplasmic details, nuclear details and color intensity were slightly better in microwave method than in routine method. [10] Similar studies by various authors showed no significant differences in the histologic quality when they compared these two protocols. [9,[11][12][13] From the perspective of the end product, microwave irradiation substantially shortens the time from specimen reception to diagnosis. ...
... This is in agreement with Rohr et al. [3] where histological slides were produced in 2-3 h using microwave irradiation. In a study done by Babu et al., [10] the turn around time for microwave tissue fixation and processing and staining was 75 min. In a study done by Henwood A, [14] the tat for microwave tissue fixation and processing was 80 min. ...
Aim and objectives:
The study aimed at assessment of microwave assisted tissue fixation, processing and staining and to determine if it can replace standard formalin fixed paraffin embedded processing in tissues of different thickness.
Materials and methods:
Specimens from buccal mucosa and gingiva was used in the study and were divided into three different thickness and was fixed, processed and stained according to conventional method and with a use of kitchen microwave oven respectively. The present study is the first of its kind where oral tissues was fixed, processed and stained with a kitchen microwave in three different thickness. The results obtained was statistically analyzed using IBM SPSS Statistics version 21.0 software.
Results:
The new technique of fixation, tissue processing and staining using a microwave employed in the present study represented a major change from conventional method and achieved significant reduction in time taken.
Conclusion:
The ease of application and speed of this technique significantly reduced turnaround time in diagnostic labs.
... Babu TM et al and Kango and Deshmukh used a domestic microwave oven in which the maximum temperature reached was 87°C and 58°C respectively. Stepwise assessment of the tissue temperature could not be recorded in this study, but the highest temperature attained was 84°C (16,21). Reagents: Depending upon their ability to absorb the microwave energy, some materials can be more microwavable than others. ...
... Apart from its use in tissue processing, it has also been found to be profoundly effectual in tissue fixation, tissue section staining, antigen preservation and electron microscopy. Studies have established that the quality of microwaveprocessed and microwave-stained slides is identical or slightly better than conventionally processed and stained slides (18,21,22,28,29,30). Also, microwave technology has immensely contributed in the successful antigen retrieval as well as enhanced immunostaining in immunohistochemical procedures (18,30,31,32), it has been a significant addition to the DNA extraction methods too (34,35,36). ...
Background
Methods of diagnostic molecular biology are routinely applied on formalin-fixed, paraffin-embedded tissues processed via conventional method. Recently, there has been a growing interest to use microwave technology in histopathology laboratories to overcome the deficiencies of the conventional processing method. Thefore, this study was aimed to compare and analyze the quality and quantity of DNA obtained from tissues processed by conventional and microwave tissue processing techniques and to further ascertain the applicability of the latter for PCR (polymerase chain reaction based research).
Methods
Thirty fresh tissues of oral squamous cell carcinoma (OSCC) were included, and each sample was cut into two equivalent halves. One tissue half was processed by conventional manual method whereas the other half was processed using a domestic microwave oven. DNA was obtained from all the tissues which were then subjected to Polymerase chain reaction (PCR) to evaluate GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) gene expression.
Results
The results revealed better DNA yield from microwave processed tissue while the quality of the DNA was alike from both the techniques.
Conclusion
On the basis of the results obtained, it can be concluded that DNA produced by microwave processed tissues was similar to that obtained by conventional processing technique in terms of quantity and quality. Thus, microwave processed tissue samples can be successfully used for further molecular studies and researches.
... Microwave processing not only reduced the processing time from 10-12 h to one hour but also doesn't use chemicals such as Xylene/Chloroform which are hazardous to our health [2]. Studies have shown that microwave processing is on par with conventional processing [3][4][5][6][7]. Xylene used as a deparaffinising agent is an excellent clearing agent but has detrimental effect on human health and known to cause hepatitis, pneumonia, depression etc [8]. ...
... There was almost no difference in the staining using conventional processing and staining versus xylene free techniques such as microwave processing and staining using LDW when adequacy for diagnosis was considered. Similar results were seen in other studies [3][4][5][6]. The only disadvantage of liquid dish wash solution is that it needed close monitoring and temperature of the solution was very important for complete deparaffinisation. ...
Introduction: Tissue processing using xylene/chloroform has been employed in histopathology reporting for the past 100 years. Microwave technique has not only reduced the processing time from 1 day to one hour but also doesn’t use xylene/chloroform and has been found to be on par with conventional processing. Xylene is expensive and detrimental to human health. The present study replaces chloroform with isopropanolol in tissue processing and xylene with liquid dish wash solution (LDW) in staining which is not only cheap but also non bio hazardous. Aim: To assess the efficacy of xylene free processing (microwave) and staining versus conventional tissueprocessing and hematoxylin and eosin are staining. Materials and methods: Two Tissue bits from 15 consecutively submitted samples at RL Jalappa Hospital after 1st Jan 2015 each of breast, cervix, lymph node, fat, thyroid, skin, alimentary tract, muscle, salivary gland, liver and kidney were taken and one was processed and stained using conventional method, other using xylene/chloroform free processing and conventional staining and other with both xylene/chloroform free processing and staining. The 495 sections were evaluated and scored by two pathologists independently for nuclear staining, cytoplasmic staining, uniformity, clarity and crispness. Results: In the samples evaluated xylene free processing and staining seems to be on par with conventional xylene processed and stained sections terms of nuclear and cytoplasmic detail, clarity and crispness. However Xylene cleared sections showed slightly better uniform staining (p value->0.05) Conclusion: Xylene/chloroform free processing and staining is not only a rapid but also safe alternative to conventional processing and staining using xylene. However more extensive studies need to be done on other types of tissues for validation
... The tissue sections from different organs were included, as shown in [Table/ Fig-1]. Tissue bits with a size of 1* 1* 0.5 cm were taken [10], which included neoplastic and non-neoplastic lesions. Those tissues which were processed by microwaves and their paired tissues which were processed by conventional methods were grouped into groups I and II respectively. ...
... It was observed that the shrinkage of the tissue bits was more pronounced with the DMWTP as compared to the CTP, but this did not affect the morphological quality of the tissues. This was consistent with the observations of other authors [10]. ...
Background:
Microwave irradiation has been tried as a replacement for the conventional tissue processing technique in histopathology laboratories for quite some time. Studies have shown that Domestic Microwave Tissue Processing (DMWTP) provides a faster delivery of the tissue sections with a morphology which is similar to that which is seen Conventional Tissue Processing (CTP). But many laboratories still confine the domestic microwave tissue processing method only to the handle selected specimens, for which urgent reports are needed. One of the probable reasons is that, understanding about the number of tissue sections which can be processed using a microwave oven at a time, with the appropriate quality, still remains unclear.
Aim:
The aim of this study was to quantitatively analyze the optimum number of samples that a domestic microwave could process at a time, as well as to qualitatively analyze the morphological outcome of those tissue sections with that of conventional processing.
Materials and methods:
This study was approved by the research and ethical committee of Sree Balaji Medical College and Hospital. A total of 135 paired tissue sections were included in the study. Ten tissue sections (which are mentioned hereafter as A10) were processed in a domestic microwave and their paired 10 tissues were processed by a conventional method. Subsequently, the number of tissues which was to be processed was increased to B15, C20, D25, E30 and F35, after ascertaining that the morphological qualities of the previously processed tissue sections were satisfactory. Sections of 4 μm thickness were taken and they were stained by the Haematoxylin and Eosin method. The slides of the tissues which were processed by the microwave method and the conventional method were randomly numbered, for a blind study, which were independently evaluated by two observers. The qualities of slides were assessed, based on 4 parameters: the cytoplasmic details, the nuclear details, the tissue architecture and the staining characteristics. The statistical analysis was done by using SPSS 15.0.
Results:
The morphological outcomes (quality) of the DMWTPs were comparable to that of the CTPs, when the sample load (quantity) in the microwave oven was up to 25 samples.
Conclusion:
Domestic microwave processing can be effectively used in laboratories with a maximum sample size of 25 samples per load. This has the advantage of being rapid, with its morphological quality being identical to that of conventional processing.
... [7,8] Microwave oven produces steady heat and increases the charge of tissue piercing maintaining the goodcalibre of the bony tissue specimen. [4,7,8,9] Altering electromagnetic fields are produced by non-ionizing radiation of microwaves which leads to the revolving of molecules which are dipolar like proteins as well as water. This produces molecular dynamics which leads to the causation of energy flux that will be continued tillradiation come to an end. ...
Background: In bony specimens, elimination of calcium is obtained by a method known as “Decalcification”. It is completed through the means of chemical agents such as acids, chelators etc that combine with ions of calcium. Decalcifying agent is used in regular conventional method where the hard tissue is placed at a room temperature (20-25°C) with modifications of the solution at orderly intervals until the final cutoff point is obtained. Usage of microwave oven for the process of decalcification is a new and fast method in contrast to the routine conventional method of decalcification. In this study, an attempt has been made to regulate and compare the conventional procedure of decalcification with decalcification done by microwave oven of hard tissue specimens by using nitric acid of 10% concentration with regards to decalcification speed, conservation of tissue architecture as well as productiveness of staining. Objectives: The study will made a comparison of Conventional and Microwave Bone Decalcification Methods by using 10% Nitric Acid. Methodology: This prospective analytical study, will include decalcification of 30 hard tissue specimens by microwave method and conventional method. The results will be compared in terms of decalcification speed, conservation of tissue architecture and staining productiveness. Necessary tests will be applied to analyze the data. Expected Results: Significant advantages of microwave method are expected over conventional method of decalcification. Conclusion: The conclusion will be drawn based on careful analysis of the results.
... The metal screen reflects the microwave and allows the inside of the cavity to be viewed while the visible light wavelengths will also travel through. The microwaves could not pass the screen, since the gaps in the screen are much smaller than the microwaves (Babu et al., 2011). So-called 'transparent microwave' materials, including some glass, pottery, paper, and many plastics, enable waves to move through. ...
This study was to evaluate the effects of microwave device in melanin pigment staining procedures and to compare between the classical Masson-Fontana staining technique at room temperature and the Microwave technique with Masson-Fontana stain. A total of 50 skin tissue samples were taken for processing. After processing, two histochemical staining techniques were carried out; the classical Masson-Fontana method at room temperature and the microwave device Masson-Fontana staining method at temperature 25°C. Both methods were compared. This study indicated that the use of a microwave device in staining of melanin pigment reduced the time significantly (P ≤ 0.001) from overnight (24 hours) to only 2 minutes with excellent staining quality as compared to the classical Masson-Fontana method (P ≤ 0.001). In conclusion, the use of a microwave device is a vital tool for staining of melanin with the Masson-Fontana method as it is a time reducer and staining quality enhancer.
... This has resulted in a substantial reduction in the basic steps of histoprocessing, thereby reducing the turn-around time and permitting same day reporting. [9] The technique of microwave-assisted tissue processing has been achieving great acceptance over the last decade, and this has led to the production of commercially available microwaves specifically designed for tissue processing; however, the cost involved in these is high. [2,10] Domestic microwaves are readily available, affordable and have been used earlier for tissue processing with good results. ...
... Cellular clarity, Colour intensity and overall Staining quality in our study was slightly higher for microwave method, in concordance with Babu et al. 4 and Nangia et al. 7 No statistically signiicant variation was observed between the two methods (p>0.05). ...
p> Background: Histopathological examination of tissues requires sliver of formalin fixed tissue that has been chemically processed and then stained with Haematoxylin and Eosin. The time honored conventional method of tissue processing, which requires 12 to 13 hours for completion, is employed at majority of laboratories but is now seeing the</p
... The development of the frozen section technique allowed a rapid processing time for intraoperative histopathologic evaluation, but this technique is associated with difficulty in preparing and cutting certain types of specimens (fatty tissues) as well as certain morphologic changes that make histopathologic interpretation more difficult. These limitations have inhibited the use of frozen sections from becoming the predominant tissue processing method for routine specimens [2]. Xylene has been traditionally used as a clearing agent in tissue processing for many years, primarily because it is miscible with both alcohol and wax while doing an excellent job of clearing the tissue of alcohol [3]. ...
ABSTRACT
Introduction: 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.
Aim: To compare conventional and microwave based decalci-fication, processing and staining of tooth and mandibular bone specimens using 5% nitric acid as decalcifying agent.
Materials and Methods: 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.
R
esults: 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.
C
onclusion: 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
Introduction:
Conventional formalin-fixed, paraffin-embedded tissue provides superior cellular morphology and long-term storage. Problems with formalin fixation comprise delay of fixation and variations in the duration of fixation. Microwave assisted tissue fixation removes the use of noxious and potentially toxic formalin that decreases the turnaround time and creates a personnel friendly workflow.
Material and methods:
The present study was conducted over a period of two years. One hundred and forty paired tissue sections were taken including both neoplastic and non-neoplastic tissues. One of the paired tissues was fixed in formalin and the other was fixed by using microwave irradiation in phosphate buffered saline. Both were then processed by conventional method. Each slide was examined and rated for the adequacy of fixation by two pathologists in a blinded fashion using 7 parameters: Cellular outline, cytoplasmic detail, nuclear detail, erythrocyte integrity, lymphocyte appearance, overall morphology and overall staining.
Results:
Statistical analysis showed that sections obtained from microwave fixed tissues were comparable to that of routinely fixed tissue. The p-values of all parameters were not significant except for the overall morphology for which p-value was significant owing to loss of tissue in some cases.
Conclusion:
Microwave irradiation substantially shortened the time from specimen reception to diagnosis (turnaround time) and allowed same-day tissue processing and diagnosis of specimens without compromising the overall quality of the histologic section.
Rapid processing of histopathologic material is becoming increasingly desirable to fulfill the needs of clinicians treating acutely ill patients. Traditional techniques for rapid processing of paraffin-embedded tissues require 4 to 5 hours, delaying treatment for some critically ill patients and requiring additional shifts of technologists in the laboratory. Microwave processing further shortens this time, allowing even more rapid histopathologic diagnosis. Few data exist comparing quality of microwave-processed tissue with that processed by more traditional techniques. We randomly selected 158 paired specimens from 111 patients. One member of the pair was processed routinely overnight, while the other was processed by the rapid microwave technique. The slides then were compared for quality of histologic preparation in a blinded fashion by 2 pathologists. Eight routinely processed specimens were judged as suboptimal, while 6 microwave-processed specimens were judged as suboptimal and 1 was considered unsatisfactory for evaluation. In the remaining cases, the material obtained by the 2 techniques was considered of identical quality. Microwave processing considerably shortens the preparation time for permanent histologic sections without a demonstrable decrease in section quality or "readability."
In this paper we describe a method of preparing tissue blocks for paraffin sections within 30 min. The method is based on microwave-stimulated diffusion reducing the dehydrating, clearing, and impregnating times by a factor of 48. The developed technique was inspired by the experimentally observed sizable temperature-dependence of viscosity and other transport properties of liquids. It is clear that, by considering the theoretical aspects of diffusion and by analysis of the influence of the used chemicals in different tissue depths, histotechnical results can be optimized. The histotechnical microwave results are light-microscopically excellent and indistinguishable from those of the well-performed 'classical' method. The nuclear size of several cell types hardly differs in both methods. The new method is valuable in particular for individual cases in which a fast diagnosis is asked for, and in which a frozen-section diagnosis is thought to be too unreliable. In addition, this method can be used in small research laboratories processing small quantities of histological material. The only equipment needed to prepare tissue blocks of optimal quality is the microwave oven.
Tissue blocks taken from healthy human lung tisues, from primary bronchial carcinoma and from mediastinal and hilar lymph nodes were placed in the following solutions. Tris buffer; buffered formalin (0.5%, 1%, 7%); 0.1 mol NaCl; distilled water; DMSO (1%, 10%, 20%); acetone (10%); methanol (50%, 80%, 100%); glutaraldehyde (2.5%), and fixed by use of a commercial microwave oven.
Tissue blocks obtained from the same surgical specimens were fixed in 7% buffered formalin for 24 h for comparison. Conventional and microwave-fixed tissue was embedded in paraffin, and stained with haematoxylin and eosin. Fifteen specimens of each group and each solution were examined by light microscopy. Minimum diameter and area of 100 nuclei of each specimen were measured interactively.
Histomorphological sections fixed with Tris buffer in a microwave oven revealed best morphological results showing more contrast in chromatin distribution of nuclei and ‘opening’ of interstitial lung capillaries in comparison to conventional formalin-fixed specimens. No statistically significant differences in area and minimum diameter of nuclei between the different groups were found. Microwave fixation using Tris buffer is a time-saving fixation method at least comparable to conventional formalin fixation. It is not accompanied by hazards to the environment that are unavoidable by formalin fixation.
This paper evaluates and extends the novel method of preparing tissue blocks for paraffin sections within 30 to 60 min, that was proposed in early 1985 in a paper by Boon et al. (1986). More than 2 years' additional experience and testing various microwave ovens has led to new protocols reported in this paper. Results are given of testing (i) an especially designed microwave oven for histoprocessing, (ii) microwavable reagents, (iii) processing larger numbers of specimens simultaneously, (iv) handling different types and sizes of tissue. It is concluded that effective temperature control offers substantial advantages. In addition, the possibilities of performing routine diagnostic pathology omitting formalin altogether are sketched.
In this article the authors describe a procedure to obtain cryostat sections of superior quality using microwave stimulation of a fixation reagent with suitable microwavability properties. In fact, the light microscopic images are of a quality that is comparable to that of the best conventional paraffin technique. The new procedure requires less time than the existing cryostat technique. The only additional piece of equipment needed in this procedure is a relatively inexpensive commercially available microwave oven.
Concerns about the toxicity of formalin dictate the need for alternative methods of tissue fixation. Alcohol-based fixatives have been proposed as optimal for immunohistochemical and nucleic acid methods and may be useful for diagnostic light microscopy. We recently converted our surgical pathology laboratory from 10% neutral buffered formalin to an alcoholic fixative (56% ethanol and 20% polyethylene glycol) and compared 100 consecutive cases after conversion with 100 formalin-fixed cases prior to conversion. One representative slide from each case was reviewed independently by four pathologists without knowledge of the fixative employed. The quality of fixation of each case was graded (0, very poor; 1, below average; 2, average; and 3, above average) based on the subjective composite evaluation of seven morphological features: tissue architecture, cell borders, cytoplasm, nuclear contours, chromatin texture, red blood cell membranes, and uniformity of staining. No significant difference in the quality of fixation was observed, although the average score of the alcoholic fixative was slightly lower than that of the formalin (2.59 vs 2.74). Interobserver concordance was 74%. Advantages of the alcoholic fixative included elimination of carcinogenic vapors, more rapid fixation, greater stain avidity, elimination of enzyme predigestion in immunohistochemical studies, and simple and rapid disposal. Disadvantages included slightly increased viscosity, increased variability of tissue staining and nuclear shrinkage, artifactual pigment deposition in bloody specimens, and increased flammability risk. Alcohol-polyethylene glycol appears to be a satisfactory alternative to formalin in routine diagnostic surgical pathology.