In April 1999, the guidance document “Histopathology reporting in cervical screening” was issued.¹ On 45 of that document is the statement “As the appearance of the tissues, even in small biopsies, often shows considerable variation, several levels are required to ensure that small foci of disease are identified”. No more specific guidance was given in the document. The issue of what constitutes “several levels” was discussed at the Symposium of Gynaecological Pathology held by the British Divison of the International Academy of Pathology in Sheffield on 10 September 1999. It was clear from the discussion that there were many varying practices being used. The practice in the laboratory at the City Hospital Nottingham was to examine two sections at each of three levels of the material, all mounted on to one glass slide. It was decided that this practice should be audited against examining two sections at each of six levels to see whether extra information was gained by this or whether important diagnostic features were being missed by using the existing practice.

The subsequent consecutive 100 colposcopic biopsies were processed according to the standard operating procedure in the laboratory and then two sections 2 μm thick were cut at each of six levels through the material. The levelling was rigorously controlled at 50 μm between each level. The levels were mounted as sections one to three on one slide and sections four to six on a second slide.

The samples were examined microscopically (all by JJ). The slide with levels one to three was examined and the diagnostic features recorded. Only then was the slide with the levels four to six examined. Any variance from the features seen in the first three levels was recorded and commented upon on the record sheet for the audit.

In only seven cases of the 100 examined was further information obtained from the second three levels (levels four to six). In four of these, the comment after examining the first three levels was that levels four to six would have been requested to be cut (always an option) because the diagnosis was not clear on the first three levels and it was felt that further sections might help to clarify the picture. A further three levels would have been requested on six cases; these four were included in the six. In the other two cases no further information was available in the extra three levels. Of the other three cases where additional information was obtained from the second three levels, two showed koilocytes in the squamous epithelium, which were not visible in the first three. This is not a clinically important finding because the management of the women would not have been affected. In the third case, the second three levels revealed focal stromal inflammation, not visible in the first three levels, also not clinically important.

Relevant histological features are almost always visible on the examination of sections from each of three levels cut from small colposcopic biopsies. In the small number of cases where the diagnosis is not clear on the first three levels, examination of the next three levels (in four of six such cases in our series) may help the pathologist to make the diagnosis. In none of our 100 cases was relevant diagnostic material missed by examining the material at three rather than at six levels. The conclusion reached has been to continue our original practice, thereby preventing the use of extra sectioning time and of twice the number of slides for each case. Critical evaluation of one versus two sections from each level was not conducted but one section would save very little cutting time and six sections sit easily on one slide.

During the course of our study, a letter appeared in the Journal of Clinical Pathology stating that sections at levels through the tissue should not be mounted on the same slide because the histological material may not be covered by the coverslip, or might be obscured by mountant.² This was not the case on any of our 200 slides. With careful placing of the material on the slide by the section cutter (fig 1) and an automated coverslipping machine the artefacts mentioned in that letter are not encountered.

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Figure 1

Three slides showing two sections at each of three levels mounted on one slide. The first level is closest to the label. The ink marks have been put on to the coverslip by the pathologist to encircle the material for examination at each level and to guide the eye from level to level when examining the material under the microscope (especially useful when the levels are offset on the slide).