1. Department of Laboratory Medicine & Pathology, Nepal Mediciti Hospital Sainbu, Nepal.
2. Department of Laboratory Medicine & Pathology, Nepal Mediciti Hospital Sainbu, Nepal.

* Correspondence: acharyashoshan6@gmail.com

Keywords: Frozen Section, Permanent Section, Accuracy, Discrepancies

1. INTRODUCTION

Frozen section plays a major role in the surgical management of patients with neoplastic and non-neoplastic disease. Since its introduction in the late 20^th century, the use of frozen section has spectacularly increased. It provides the surgeon with important pathologic information while the patient is on the operating table [1]. There are many indications for performing frozen section, including determination of the nature and extent of a lesion, evaluation of surgical margins and assessment of adequacy of tissue for diagnosis [2]. However, the main purpose of frozen section is to guide the surgeon making immediate decision as the extent and or adequacy of surgical procedure, thus decreasing the need for reoperation [3]. The surgeon’s confidence in frozen section results, depends on the diagnostic accuracy of this procedure [4]. Frozen section diagnosis is usually compared to permanent section diagnosis to evaluate the diagnostic accuracy. Evaluating discrepancies, identifying deficiencies and resolving the underlying problems can improve the accuracy of frozen section [5]. Reasons for diagnostic discrepancy generally fall into one of the following categories: technical problems, sampling errors or interpretation errors [6]. It is well established that periodic review of frozen sections result in improvement of performance. Therefore, it has been suggested to place periodic review as part of quality assurance in pathology departments [7]. The use of intra-operative frozen section to distinguish between benign and malignant histology is reliable in guiding operative decision-making to determine the extent of surgical staging required [8]. Frozen section was first performed by Welch in 1891 for intra-operative consultation and further development was contributed by Cullen in 1895 and Wilson in 1905 [9].The cryostat that contain rotary microtome that maintain inside temperature of the cryostat about −20° to −50° Celsius. OCT (optimal cutting temperature) medium is used to quickly embed fresh tissue specimens for frozen section using cryostat.  Performing a Frozen section analysis is a multistep process which begins with retrieval of the specimen from patient by surgeon which is followed by preparation of slides and microscopic analysis by pathologist, possible specimen triage for further workup, and finally rendering the FS diagnosis [10]. FS diagnosis is usually compared to permanent section diagnosis to evaluate the diagnostic accuracy. Compared to other intra-operative diagnostic methods, the tissue structure is better preserved in FS analysis and this method better visualize the structural arrangement of cells, stroma and the tumor [11].   Evaluating discrepancies and resolving the underlying problems we can improve the accuracy of frozen section [12]. However, frozen section has some limitations like artifacts, poor quality of sections, suboptimal histology, poor quality staining, and pressure of immediate diagnosis and lack of ancillary studies at the time of intraoperative consultation. Therefore frozen sections cannot be considered as replacement of the permanent sections [13]. Discussion of the cases is important before frozen section, case details including relevant clinical and radiological information should be provided to the reporting pathologist. The surgeon and the pathologist should act like a team to achieve a meaningful frozen report.  Diagnostic discrepancy reasons generally fall into one of the following categories, technical problems, sampling errors or interpretation errors [14]. Frozen section had to undergo periodic review that result in improvement of performance and quality assurance in pathology department. Therefore, it has been suggested to place periodic review as part of quality assurance in pathology departments [15]. In the result of extensive study conducted by the College of American Pathologists (CAP) on more than 90,000 diagnoses made through frozen section analysis in 461 different medical centers, the diagnostic accuracy result of frozen section was 98.52% [16]. The study reviewed and analyzed frozen sections performed in our institute during a 2 year period to assess the diagnostic accuracy and to determine the rates of deferred and discordant results as well as the reasons for discrepancy.

2. MATERIALS & METHODS

It retrospectively analyzed the frozen section cases performed in Pathology Department, Nepal Mediciti Hospital, for a period of 2 years. It has been chosen the period because the electronic data was available well in this period and sample size in this period was more than other period. Nepal mediciti hospital is dedicated to assure world class health service with finest technology, infrastructure with class A laboratory, with NABH (National accreditation board for hospitals and healthcare) certified. Fresh tissue was sent in a clean plastic container without any fixative or saline (to avoid fixation artifacts/degenerative changes) along with requisition form with complete clinical details from the operation theater. Indication of frozen section was recorded. Patient files in the pathology department provided data regarding the frozen section cases. Tissue specimens sent for frozen section were frozen and cut by a cryostat machine, for frozen section gross examination was done, specimen dissected and sections were taken from representative areas. Frozen sectioning was done on SLEE cryostat Mainz. Tissue was kept in the company’s tissue embedding medium and placed in object disc inside the cryostat and was frozen. The cryostat was set at a range between -26°C to -38°C depending upon the nature of the tissue. When the medium and tissue were frozen, the chuck was inserted into the clamping lever and was fixed. Clearance angle was set at 12°C. Sections were cut at a thickness of 5-8 μ and were immediately fixed in 95% isopropyl alcohol. Rapid hematoxylin and eosin (H&E) staining was performed. Frozen section diagnosis was made by consensus of at least two pathologists, including one senior pathologist in correlation with appropriate clinical details. In presence of inter observer variation between two pathologist, the section was consulted with the third or even more pathologist. The diagnosis made by the majority is considered the final diagnosis. The final diagnosis was immediately conveyed to the operating surgeon. The turnaround time of entire procedure from receipt of specimen to delivery of report was recorded. The remaining tissues if available were fixed in 10% formalin, grossed and adequate representative sections were taken according to the standard guidelines. The sections were then evaluated in H&E stain. The frozen section diagnoses were compared to that of the permanent sections, to assess the accuracy of the technique. The frozen section results in comparison to final diagnoses were then categorized into three groups, concordant, discordant and deferred. Diagnosis was considered as concordant if there was agreement and discordant if there was disagreement with permanent section diagnoses. Deferred cases were defined as indeterminate diagnoses at the time of frozen section examination. Deferral rate was not included in the calculation of accuracy. Finally, discordant cases were reviewed and causes of discrepancy were recorded. The electronic data system and record files in the pathology department were used to get data regarding the frozen section. The frozen section diagnoses were compared to that of the permanent sections, to assess the accuracy of the technique. The results that we got from frozen section were then compared to final diagnoses and results were then categorized into three groups i.e. concordant, discordant and deferred. After analyzing the results of proper frozen section study, diagnoses were categorized as concordant if there was agreement with permanent section diagnosis and discordant if there was disagreement with permanent section diagnoses. Deferred cases were defined as indeterminate diagnoses at the time of frozen section examination. While calculating the accuracy deferral rate was not included. Finally, discordant cases were reviewed and causes of discrepancy were recorded.

3. RESULTS & DISCUSSIONS

In the 2 year period of our study, 100 frozen section specimens were received from surgical cases. The submitted tissues for frozen section were primarily from hepatobiliary then uterus, ovary, head and neck and breast respectively (Figure 01). Indications for frozen section were (1) verification and categorization of neoplasms in 85 (85%), (2) evaluation of margins in 12 (12. %) and, (3) determination of the organ of origin 3 (3.0%) of cases.

Figure 01: Frozen section from hepatobiliary, uterus, ovary, head, neck and breast

In total 100 cases, in 4 (2.0 %) cases the diagnosis of frozen section was deferred to permanent section. Among remaining 96 cases, frozen section and permanent section diagnoses were concordant in 94 (94.0%) and discordant in 4 (4.0%) of cases (Table 01). In our study, there were no false-positive or inconsistent cases. The four discordant case were in the category of false-negative and the rest were divided between true positive and true negative cases. Sensitivity was calculated by TP/TP+FN. Specificity was calculated by TN/TN+FP

Table 01: Frequency of concordant, discordant and deferral cases

Among discordant cases, one is from hepatobiliary, one from ovary, one from head and neck and one from others. We reviewed the cases of frozen section slide to assess the reason for discrepancy, (Table 02). Among 4 discordant cases the reason of discrepancy was sampling error in two (50 %) case and misinterpretation in two (50 %). Sensitivity, specificity, positive predictive value and negative predictive value of frozen section in comparison with permanent section were 100 %, 96.6 %, 100 % and 96 .6 % respectively.

Table 02: Discordant cases

4. DISCUSSION

The frozen section is a rapid diagnostic method to assist the surgeon in intraoperative management of surgical procedures by avoiding second surgery [17]. It is costly and requires skill in technique as well as interpretation. Frozen section has some limitations such as freezing artifacts, sampling, section cutting, staining quality, and interpretation so it cannot replace paraffin embedded tissue technique though it has benefit that it provides rapid diagnosis [18]. Frozen section should always be followed by permanent section as permanent section has benefit for performing special stain and Immunohistochemistry.  In the study we conducted we reviewed the frozen sections performed in Pathology Department of Nepal Mediciti Hospital for 2 year period to evaluate the diagnostic accuracy of the test in this institution. We analyzed the study which revealed an accuracy rate of 100% in benign lesions and 96% in malignancy. Freezing artifacts were noted in some sections, which were re- evaluated. The false-positive rate (0%) and false-negative rate (2 %) were well within the range reported in the literature. The discordant cases were reviewed then to find the reasons for discrepancy. In our study total number of concordant and discordant cases were 94 (94.0%) and 4 (4.0%) respectively. Deferred rate is also a valid parameter of quality assurance. Our study showed 2 (2.0 %) deferral cases which is comparable to previously published studies with a deferred rate ranging from 0.04% to 6.7% [19].  Deferral rates in frozen section may vary according to clinical expertise and also clinical setting of pathology department and the type of specimens encountered [20]. One of the study evaluated the accuracy of frozen section in 847 consecutive specimens in a teaching hospital. Their results showed concordant, discordant and deferred rates of 92.6%, 1.7% and 5.8% respectively which is comparable with our results [21]. The previously published studies in which discordant rates were studied have shown result vary in discordant rates from 1.4 to 11.8 percent, which is comparable` with our result 4 percent. Concordance and discordance between frozen section and histopathology may differ among lesions and malignant conditions. A study showed the concordant rate of 83% in gallbladder, 100% in GI tract tissues, ovarian tissues, and oral cavity tissues, similarly another study showed the concordant rates of 100% in GI tract, oral cavity and soft tissue samples, whereas 90.9% concordance in ovarian and 93.75% in Breast tissues. The results are similar to this study [22], other previous studies show misinterpretation as the main cause for discrepancy followed by sampling error. However, it showed equal number of cases of misinterpretation and sampling error. In the present study discordant rate is slightly higher than previously published data. This may be because of the lack of highly experienced or trained pathologist in the institution. Our results show all false negative cases and no false positive cases. This result is comparable to previous studies [23], there is a lower deferred and error rate when specimens are interpreted by more experienced pathologists. A study has shown junior residents make higher percentage of inaccurate diagnoses which is improved with additional training [24]. Evaluation of the specimens by two observers or even three, when there is uncertainty, reduces the rate of error [25]. In our study diagnoses on frozen sections made by residents was reviewed by attending two or three pathologists. A study has shown junior residents make higher percentage of inaccurate diagnoses which is improved with additional training. In the review, author included a total of 37 studies, which were mostly from united state found that FS pathology to have a pooled sensitivity and specificity of 83 ± 13% and 95 ± 8%. Our reported sensitivity and specificity were also similar to the mentioned reports [26]. As per The College of American Pathologists (CAP) cancer protocols the turnaround time in frozen section should be within 20 minutes. Turnaround time does not include transport time prior to receipt of specimen, and also excludes cases where multiple sequential studies are performed [27]. The average turn-around time observed in the present study was 20 minutes, which is comparable with the range reported by other studies [28]. The frequent limitation observed in present study were technical errors sampling errors, sectioning, inadequate history, staining and labeling. An important technical factor causing difficulty in interpretation is quality of section which limits the evaluation of cellular details, such limitations are seen in other studies as well [29,30].

5. CONCLUSIONS

Frozen section is an accurate and reliable test when accurate sampling plus better communication between pathologist and surgeon is made which then helps in reducing the rate of discordant and deferral cases. Reevaluation of the interpretations by a second pathologist especially when there is uncertainty is helpful in reducing the diagnostic errors. The intra-operative consultation using FS is a very useful but one needs to be aware of its indication and limitations. Methodological gross/ macroscopic examination, accurate sampling, avoiding technical errors in sectioning and staining, and better communication between pathologist and surgeon along with reevaluation of the interpretations by a second or may be third experienced pathologist is helpful in reducing discordant and deferral rates. This provides rapid, reliable and cost effective details necessary for rapid diagnosis and on table patient management.

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