Histologic processing artifacts and inter-pathologist variation in measurement of inked margins of canine mast cell tumors

Abstract

Although quantitative assessment of margins is recommended for describing excision of cutaneous malignancies, there is poor understanding of limitations associated with this technique. We described and quantified histologic artifacts in inked margins and determined the association between artifacts and variance in histologic tumor-free margin (HTFM) measurements based on a novel grading scheme applied to 50 sections of normal canine skin and 56 radial margins taken from 15 different canine mast cell tumors (MCTs). Three broad categories of artifact were 1) tissue deformation at inked edges, 2) ink-associated artifacts, and 3) sectioning-associated artifacts. The most common artifacts in MCT margins were ink-associated artifacts, specifically ink absent from an edge (mean prevalence: 50%) and inappropriate ink coloring (mean: 45%). The prevalence of other artifacts in MCT skin was 4-50%. In MCT margins, frequency-adjusted kappa statistics found fair or better inter-rater reliability for 9 of 10 artifacts; intra-rater reliability was moderate or better in 9 of 10 artifacts. Digital HTFM measurements by 5 blinded pathologists had a median standard deviation (SD) of 1.9 mm (interquartile range: 0.8-3.6 mm; range: 0-6.2 mm). Intraclass correlation coefficients demonstrated good inter-pathologist reliability in HTFM measurement (κ = 0.81). Spearman rank correlation coefficients found negligible correlation between artifacts and HTFM SDs ( r ≤ 0.3). These data confirm that although histologic artifacts commonly occur in inked margin specimens, artifacts are not meaningfully associated with variation in HTFM measurements. Investigators can use the grading scheme presented herein to identify artifacts associated with tissue processing.

Evaluation of a modified proportional margin approach for complete surgical excision of canine cutaneous mast cell tumours and its association with clinical outcome

“…Following surgical excision of cutaneous masses and submission of samples through fixation in formalin, there are variable degrees of tissue shrinkage, with the possibility of ink-associated artefacts including ink being absent from an edge as well as inappropriate ink colouring also confounding HTFM assessment. 18,[22][23] Given the subjectivity of HTFM measurement, there may also be inter-observer variation. 22 As a result, measurement of histologic margins will invariably contain a degree of error.…”

“…22 As a result, measurement of histologic margins will invariably contain a degree of error. Kiser et al 23 attempted to quantify histologic processing artefacts and inter-pathologist variation in the measurement of the HTFM in canine MCTs. They noted there was good inter-pathologist reliability with HTFM measurement (interobserver median SD of 1.9 mm); however, there was a common incidence of artefacts including tissue deformation at inked edges, inkassociated artefacts and sectioning-associated artefacts.…”

Histologic margins and the residual tumour classification scheme: Is it time to use a validated scheme in human oncology to standardise margin assessment in veterinary oncology?

“…The application of ink to the surgical resection margins is recommended to orientate the pathologist and provide accurate histologic identification of the true surgical margins . Purple or red coloured inks are not recommended because the surgical ink is difficult to differentiate from the colours of haematoxylin and eosin stains . In one study in which a consistent ink application method was used, the ink dissected along fascial planes in 28.1% of samples and inadvertently adhered to surfaces other than the surgical margin in 68.1% of samples .…”

“…26 Purple or red coloured inks are not recommended because the surgical ink is difficult to differentiate from the colours of haematoxylin and eosin stains. 59,60 In one study in which a consistent ink application method was used, the ink dissected along fascial planes in 28.1% of samples and inadvertently adhered to surfaces other than the surgical margin in 68.1% of samples. 4 While inking has the potential to confound the interpretation of histologic margins, the application of ink to the lateral and deep surgical margins is essential for margin assessment and should be performed routinely using a technique which minimises these potentially confusing artefacts.…”

Assessment of a computerized quantitative quality control tool for whole slide images of kidney biopsies

“…This delay is usually caused by poor‐quality glass slides needing to be reproduced or WSIs rescanned and adds unnecessarily to storage overheads if the slides are not diagnostically usable. Evidence suggests that although clinical pathologists’ interpretations are not impacted by differences in WSI quality [2], these differences may negatively affect the performance of digital pathology‐based computational tools, including machine or deep learning algorithms [3,4]. Current best practices in digital pathology include the manual quality control (QC) of WSIs before experimental execution.…”