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 Table of Contents  
ORIGINAL ARTICLE
Year : 2016  |  Volume : 3  |  Issue : 2  |  Page : 57-62

Evaluation of diagnostic utility of step sections in dermatopathology: A prospective study of 200 consecutive punch biopsies


1 Jerath Path Labs, Jalandhar, Punjab, India
2 Department of Pathology, Government Medical College and Hospital, Chandigarh, India
3 Division of Pathology, National Institute for Research in Environmental Health, Bhopal, Madhya Pradesh, India
4 Department of Dermatology and Venerology, Government Medical College and Hospital, Chandigarh, India

Date of Web Publication5-Dec-2016

Correspondence Address:
Ujjawal Khurana
Flat No. 4017, Type IV Flats, AIIMS Residential Campus, Bhopal, Madhya Pradesh
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/2349-6029.195219

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  Abstract 


Background: Obtaining deeper sections or step sections is a common practice for small skin biopsies. Much of the available literature highlights the importance of step sections in neoplastic diseases of skin. However, the routine dermatopathology practice in developing countries shows a predominant burden of nonneoplastic diseases, and the utility of step sections in this context has not been much reported. Objective: The study was aimed to evaluate the utility of prospective step sections in routine dermatopathology practice. Materials and Methods: The present study comprising 200 consecutive skin biopsies was carried out in a prospective manner. Three slides were prepared in each case: Slide 0 was prepared from the ribbon of tissue obtained from untrimmed block, step sections 1 and 2 were obtained at 50 μm and 100 μm depth, respectively. The diagnosis was rendered on slide 0 and subsequently reviewed after examining step section 1 and 2. Results: Of the 200 cases, additional findings on step sections were found in 18 cases (9%) which led to change in diagnosis in 10 (5%) cases. Step section 1 led to correct diagnosis in 6 cases (3%). Step section 2 led to correct diagnosis in 10 cases (5%); however, this was statistically not significant (P ≥ 0.065) when comparing to step section 1. Additional findings which led to diagnosis was most commonly found in the cases of borderline tuberculoid leprosy (5 out of 10 cases) followed by bullous disorders. Conclusion: We therefore believe that step sections improve the diagnostic accuracy in skin biopsies and they are, especially useful in suspected cases of Hansen's disease. There is no statistical advantage of step section 2 versus step section 1; although, step section 2 had shown to include all the additional findings which led to a change in diagnosis.

Keywords: Leprosy, prospective, skin biopsy, step sections


How to cite this article:
Jerath P, Punia R, Khurana U, Thami G P, Handa U, Mohan H. Evaluation of diagnostic utility of step sections in dermatopathology: A prospective study of 200 consecutive punch biopsies. Indian J Dermatopathol Diagn Dermatol 2016;3:57-62

How to cite this URL:
Jerath P, Punia R, Khurana U, Thami G P, Handa U, Mohan H. Evaluation of diagnostic utility of step sections in dermatopathology: A prospective study of 200 consecutive punch biopsies. Indian J Dermatopathol Diagn Dermatol [serial online] 2016 [cited 2020 Oct 28];3:57-62. Available from: https://www.ijdpdd.com/text.asp?2016/3/2/57/195219




  Introduction Top


Skin biopsy is indicated in all suspected dermatoses and to clarify a diagnosis when a number of cutaneous entities are under consideration.[1] Incomplete sectioning through the tissue block is a known cause of false-negative diagnosis in dermatopathology.[2],[3] Deeper sectioning of small biopsy specimens such as needle core and endoscopic biopsies is commonly used to enhance diagnostic sensitivity and accuracy. Deeper sections in biopsies can be random or in form of step sections. Step sections are taken at a fixed distance, usually 50 µm from each other and subsequently labeled as step one, two, three, etc. The utility of step sections has been evaluated in prostatic, rectal, oral, colonic, esophageal, trephine, and sentinel lymph node biopsies.[4],[5],[6],[7],[8],[9]

In some laboratories, step sections or “deeper levels” are prepared before receipt of the slides by the pathologist (prospective step sections), and in some laboratories, they are requested after reviewing the initial slide.[2],[10] The use of prospective step sections may improve diagnostic accuracy and reduce the time for diagnosis.

The present study was carried out with the aim to evaluate the diagnostic utility of prospective step sections and to compare the diagnostic utility of step section 1 in comparison to step section 2.


  Materials and Methods Top


The present study was conducted on consecutive 200 punch skin biopsies received in the Department of Pathology from the Dermatology department irrespective of the provisional diagnosis and type of biopsy. The study excluded excisions. The punch biopsies received were of 3.5 mm, that is, 3500 µm in thickness. In case of patch, plaque, and macular lesions, the punch was taken from the center of lesion. In vesiculobullous/papular disorders, early intact vesicles/bulla/papule with scanty normal margin was preferred. The punch biopsies were submitted intact. All the skin biopsies comprising the study were received in formalin and had both dermis and epidermis on microscopic examination.

The biopsies underwent routine tissue processing and paraffin embedding. Sections of 3 µm thickness were prepared from the paraffin blocks. For each specimen, 3 prospective sections, each consisting of 1 ribbon of tissue containing 4–6 sections, were obtained at 50 μm intervals from the paraffin block. The initial slide was prepared from the ribbon of tissue obtained from untrimmed block of tissue and was labeled as slide 0; the second slide obtained after 50 μm interval was labeled as step section 1 and the third slide obtained after 50 μm interval was labeled as step section 2. Sections were stained with hematoxylin and eosin. Special histochemical (periodic acid-Schiff [PAS], Masson's trichrome, Elastic Van Gieson, Lepra, acid-fast bacteria [AFB]) and immunohistochemical (IHC) stains were done wherever required. Sections for special stains and IHC were taken after sections for slide 0 had been taken if the orders had been placed at the time of grossing or after SS2 if the orders had been placed subsequently.

A diagnosis was rendered using slide 0 only, and subsequently, step section 1 and step section 2 were reviewed. The utility of step sections were classified into:

  • No additional information
  • Additional findings which contributed to the diagnosis rendered earlier on slide 0
  • Additional findings which led to diagnosis or changed the diagnosis.


Statistical analysis was performed using Chi-square test and simple proportion test.


  Results Top


These 200 biopsies were received over approximately 1 year 3 months. Out of a total of 7114 surgical pathology specimens received over this period, the burden of skin punch biopsy was 2.8% (200/7114). The patients' age ranged from 1 to 80 years with a mean of 37.9 ± 18.19 years. Maximum number of cases were in the third decade of life. Of 200 cases, 119 were males and 81 were females with a ratio of 1.4:1. The most common clinical finding was a patch or a plaque (43 cases each = 21.5%), followed by macular or ulcerated lesion (35 cases each = 17.5%), followed by papular type of lesion (18 = 9%). The most common provisional clinical diagnosis was Hansen's disease followed by cutaneous vasculitis. Out of 200 cases, 191 (95.5%) cases were reported as nonneoplastic/benign and 9 (4.5%) as malignant [Table 1] and [Table 2].
Table 1: Spectrum of nonneoplastic lesions and benign tumors observed in the study (n=191)

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Table 2: Spectrum of malignant lesions observed in the study (n=9)

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The special stains (Lepra, AFB, PAS, Masson's Trichrome, Elastic Van Gieson) were required in 76 out of 200 cases (38%). Some cases required only one special stain, but in some cases, more than one special stain was required for diagnostic workup. In cases suspected of Hansen's, an order for lepra stain was put at the time of grossing. Once the special stain had been carried out it was not repeated, only in the cases where the section had floated or stain had not worked properly a repeat stain was requested.

On step section 1 and 2 combined together, additional findings were observed in 18 cases (9%). Thus, in 9% of cases, step sections were helpful. The point estimate is 9%, and the preferred (Wilson score) 95% confidence limits are 5.76%–13.78%.

In 10 cases, additional findings led to diagnosis or changed the diagnosis, whereas in 8 cases, the diagnosis was not changed but only contributed to the diagnosis already made on slide 0.

Additional findings which led to diagnosis or changed the diagnosis (10 cases)

In 5 cases, granulomas not present in initial slide 0 were seen in step sections 1 and/or 2 which led to a diagnosis of borderline tuberculoid (BT) leprosy [Figure 1]. There was one case each of pemphigus vulgaris [Figure 2] and bullous pemphigoid [Figure 3] in which bulla and its contents were not present in slide 0 but were present in step sections 1 and 2, and that helped in making the diagnosis. In one case of pustular psoriasis, step sections showed the presence of pustules, besides features of psoriasis leading to diagnosis. In a case of lichen planus pigmentosus (LPP), band-like infiltrate and pigment incontinence not present on the initial slide was seen on step sections. The step sections changed diagnosis from chronic nonspecific dermatitis to LPP. In yet another case of chronic folliculitis, follicular destruction not seen on initial section was present on step section leading to diagnosis. In these 10 cases, had there been no step sections the diagnosis would not have been made or diagnosis would have been wrong.
Figure 1: Photomicrograph of a case of borderline tuberculoid leprosy which was diagnosed on step sections. No granulomas were seen on slide 0 but they were seen on step sections (H and E; ×100)

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Figure 2: In this case of pemphigus vulgaris subsequent step sections showed the bulla as well as acantholytic cells which lead to the diagnosis (H and E; ×100)

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Figure 3: The slide 0 shows intraepidermal spongiosis and vesiculation; however, the step section 2 showed dermoepidermal bulla with presence of eosinophils which lead to diagnosis of bullous pemphigoid (H and E; ×100)

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Additional findings which contributed to the diagnosis rendered on slide 0 (8 cases)

Three cases were of BT leprosy, 2 showed the presence of nerve destruction which was not present in slide 0 and in one case granulomas became more in number on step sections 1 and/or 2 [Figure 4]. In a case of spongiotic dermatitis, step sections showed an additional finding of vesicle formation which was not seen on slide 0. In one case of suspected morphea, presence of adequate adenexa on serial sections ruled out the possibility of morphea. In one suspected case of dermoid cyst, only keratinous debris was seen on initial section, but the presence of adnexal structures in cyst wall in step sections helped to contribute to the diagnosis but did not change the diagnosis. In one case, each of lupus vulgaris and lupus miliaris disseminated fascei, granulomas, and caseation necrosis became more prominent, respectively, on step sections.
Figure 4: Photomicrograph of a case of Borderline tuberculoid leprosy. The granulomas became more prominent on the step sections (H and E; ×40)

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In the present study, additional findings were found in 10 cases of granulomatous inflammation, followed by 4 cases of bullous disorders, 3 cases of follicular disorders, and one case of LPP. The number of additional findings were statistically significant (P < 0.001). Step section 1 detected additional findings in 9 (4.5%) cases, whereas step section 2 detected additional findings in 16 (8%) cases out of 200 cases. On comparing step section 1 with 2, in 7 (3.5%) cases, additional findings were only seen on step section 2; however, this was statistically not significant (P ≥ 0.05) when comparing to step section 1. The diagnosis was changed in 10 out of these 18 cases. This was statistically significant (P < 0.002) compared to slide 0. Step sections 1 and 2 detected change in diagnosis in 6 (3%) and 10 (5%) cases, respectively. There was a significant overlap of six cases. On comparing step section 1 with 2, in 4 (2%) cases, change in diagnosis was seen only with step section 2; however, this was statistically not significant (P ≥ 0.065) when compared to step section 1 [Table 3].
Table 3: A brief statistical overview of the step sections in the current study

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Out of 200 cases, 9 cases were reported as malignant and remaining 191 were labeled as nonneoplastic or benign. In malignant cases, changes or additional findings were not observed in step sections. The statistical advantage of step sections in benign over malignant conditions was statistically insignificant P = 0.066 (P ≥ 0.05).


  Discussion Top


Skin biopsy material should always be examined carefully and thoroughly as incomplete sectioning through the tissue block is a known cause of false diagnosis.[2],[3],[11] In the present study, additional findings were seen in 18 (9%) cases on step section 1 and 2 combined together which led to improved diagnostic accuracy in 10 (5%) cases. In a study by Bruecks et al., prospective step sections improved diagnostic accuracy in 7% of small skin biopsies and step sections were found to be helpful in 30% of cases.[2] The study design in this study was similar to ours, that is, the inclusion of all skin biopsy cases irrespective of their clinical diagnosis. However, in a study by Maingi and Helm et al., where step sections were requested after reviewing the initial slide, the deeper section requirement has been reported to be 7%. This led to additional diagnostic information in 37.3% of the cases.[10]

The majority of cases (95%) in the present study were nonneoplastic as excisional biopsies were excluded, which is the preferred technique for the evaluation of neoplastic lesions. The additional step sections in case of malignant disorders only confirmed the diagnosis rendered on slide 0 and did not yield any additional information. As the number of malignant cases was only 9, the applicability of step sections for malignant disorders cannot be concluded. However, various authors have found the usefulness of step sections in malignancies.[11],[12],[13],[14] In cases where malignancy is strongly suspected or an in situ lesion is seen up to 10 step sections have also been taken.

In a study done on cases showing actinic keratosis in the initial level; step sections showed additional findings in 33% of cases.[12] These additional findings were malignant lesions such as squamous cell carcinoma (in situ and invasive type); basal cell carcinoma. However, additional findings in benign lesion such as seborrheic keratosis and porokeratosis were also seen. There was one case each of actinic keratosis and Bowen's disease in our study which did not show any significant finding on step sections. In another study performed on cases diagnosed as melanoma on initial level; step sections were found to be prognostically important, as variables such as tumor thickness and tumor depth were found to be greater on step sections leading to change in management in some cases.[13]

In this study, additional findings which led to diagnosis were most commonly found in BT leprosy (5 out of 10 cases) followed by bullous disorder. Step sections may help in showing the granuloma, may increase the number of granulomas, suggesting their distribution or condition of nerves. Furthermore, step sections are helpful in diagnosing leprosy reactions. Hence, in a patient with a presumptive clinical diagnosis of leprosy, we suggest prospective step sections in addition to lepra stain should be taken. Step sections should also be done in bullous disorders literature reveals that one of the bullous disorder that maximally benefits in arriving at a correct diagnosis by step sections is herpes simplex virus infection.[15] In a study from Nepal also it was suggested that a 3 mm punch with step sections may yield additional diagnostic information in granulomatous disorders and inflammatory/infective dermatitis.[16] The other nonneoplastic diseases in which step sections have been found to be useful are scabies, folliculitis, granuloma annulare, viral wart, solar lentigo, etc.[2],[10]

On comparing step section 1 and 2, step section 2 detected additional findings in 16 (8%) whereas step section 1 detected additional findings in 9 (4.5%). The difference of additional diagnosis on step sections was 7 cases (3.5%) which was statistically not significant (P ≥ 0.05). This difference as reported by Bruecks et al. was 2%.

The prospective cross-sectional studies have been used to evaluate turn around time,.[16] Prospective sections help in reducing the turn around time, but they incur a slight increase in the cost due to material cost (storage space, slides, and stains) and labor cost. However, the technicians find it more convenient to cut prospective sections because retrieval of the block, bringing it to the right temperature and then cutting it makes it more laborious and time-consuming. The retrospective sections increase the turnaround time by 1–2 days, but they are cost-effective.[2],[16] The comparison of our study with other studies available in literature is mentioned in [Table 4].
Table 4: The present study is reviewed along with the studies available in literature

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  Conclusion Top


To conclude, step sections improve the diagnostic accuracy in skin biopsies especially in Hansen's disease. The authors strongly advocate the use of prospective step sections in punch biopsies; at least one step section should be taken (keeping the cost issues in mind), preferably at 100 µ, which was shown to include all the additional findings that led to the diagnosis.

Acknowledgments

The technical staff of Department of Pathology, GMCH, Chandigarh.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
  References Top

1.
Pariser RJ. Skin biopsy: Lesion selection and optimal technique. Mod Med 1989;57:82-90.  Back to cited text no. 1
    
2.
Bruecks AK, Shupe JM, Trotter MJ. Prospective step sections for small skin biopsies. Arch Pathol Lab Med 2007;131:107-11.  Back to cited text no. 2
    
3.
Penneys NS. Histopathologist: To step section or not? Arch Dermatol 2001;137:375-7.  Back to cited text no. 3
    
4.
Byar DP, Mostofi FK. Carcinoma of the prostate: Prognostic evaluation of certain pathologic features in 208 radical prostatectomies. Examined by the step-section technique. Cancer 1972;30:5-13.  Back to cited text no. 4
    
5.
Manyam R, Reena K, Saraswathi TR. Role of deeper sections in diagnostic oral histopathology: A retrospective study. Indian J Dent Res 2011;22:62-5.  Back to cited text no. 5
[PUBMED]  Medknow Journal  
6.
Surawicz CM, Meisel JL, Ylvisaker T, Saunders DR, Rubin CE. Rectal biopsy in the diagnosis of Crohn's disease: Value of multiple biopsies and serial sectioning. Gastroenterology 1981;80:66-71.  Back to cited text no. 6
    
7.
Chitkara YK, Eyre CL. Evaluation of initial and deeper sections of esophageal biopsy specimens for detection of intestinal metaplasia. Am J Clin Pathol 2005;123:886-8.  Back to cited text no. 7
    
8.
Campbell JK, Matthews JP, Seymour JF, Wolf MM, Juneja SK; Australasian Leukaemia Lymphoma Group. Optimum trephine length in the assessment of bone marrow involvement in patients with diffuse large cell lymphoma. Ann Oncol 2003;14:273-6.  Back to cited text no. 8
    
9.
Groen RS, Oosterhuis AW, Boers JE. Pathologic examination of sentinel lymph nodes in breast cancer by a single haematoxylin-eosin slide versus serial sectioning and immunocytokeratin staining: Clinical implications. Breast Cancer Res Treat 2007;105:1-5.  Back to cited text no. 9
    
10.
Maingi CP, Helm KF. Utility of deeper sections and special stains for dermatopathology specimens. J Cutan Pathol 1998;25:171-5.  Back to cited text no. 10
    
11.
Guillén DR, Cockerell CJ. Accurate diagnosis of cutaneous keratinocytic neoplasms: The importance of histological step sections (and other factors) Arch Dermatol 2000;136:535-7.  Back to cited text no. 11
    
12.
Carag HR, Prieto VG, Yballe LS, Shea CR. Utility of step sections: Demonstration of additional pathological findings in biopsy samples initially diagnosed as actinic keratosis. Arch Dermatol 2000;136:471-5.  Back to cited text no. 12
    
13.
Dyson SW, Bass J, Pomeranz J, Jaworsky C, Sigel J, Somach S. Impact of thorough block sampling in the histologic evaluation of melanomas. Arch Dermatol 2005;141:734-6.  Back to cited text no. 13
    
14.
Werner B, Mulinari-Brenner F. Saved by step sections: An unusual presentation of basal cell carcinoma. Dermatol Pract Concept 2011;1:49-52.  Back to cited text no. 14
    
15.
Resnik KS, DiLeonardo M. Herpes incognito. Am J Dermatopathol 2000;22:144-50.  Back to cited text no. 15
    
16.
Kattel G, Sinha AK, Agrawal S. Utility of prospective step sections in diagnostic skin histopathology for small biopsies. J Pathol Nepal 2014;4:552-9.  Back to cited text no. 16
    


    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4]
 
 
    Tables

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



 

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