Immunocytochemistry of effusion fluids: Introduction to SCIP approach

Abstract

Due to the remarkably wide morphologic spectrum of reactive mesothelial cells, some of the effusion fluids may be difficult to interpret with objective certainty by cytomorphology alone. Cytomorphology of well to moderately differentiated adenocarcinomas (responsible for the bulk of malignant effusions) may overlap with floridly reactive mesothelial cells. Even mesotheliomas including diffuse malignant epithelioid mesothelioma, are usually cytomorphologically bland without unequivocal features of malignancy. The intensity of challenge depends on the interpreter’s training or experience level, institutional demographics of patients (such as type of prevalent diseases, predominant sex and age group), technical support, and quality of cytopreparatory processing. In general immunocytochemistry is valuable adjunct to facilitate objective interpretation with or without other ancillary techniques as indicated. An increasing number of immunomarkers is further refining the contribution of immunohistochemistry to this field. However, application of immunohistochemistry to effusion fluids is relatively challenging because of many variables. Multiple factors such as delay after specimen collection, specimen processing related factors including fixation and storage; ambient conditions under which paraffin blocks are archived (for retrospective testing); antigen retrieval method; duration of antigen retrieval step; antibody clone and dilution; and antibody application time are identical to application of immunohistochemistry in other areas. The significant challenge related to the potential compromization of the immunoreactivity pattern due to exposure to non-formalin fixatives / reagents is also applicable to effusion fluid specimens. The immunoreactivity results would be compared and corelated with cumulative metadata based on the reported studies performed and validated on formalin-fixed paraffin-embedded tissue sections. Deviating from such protocols may lead to suboptimal results, which is not uncommon in clinical practice with potential compromization of patient care and related liability. Because of this, it is critical to perform immunocytochemistry on formalin-fixed cell-block sections only. In addition, unless the interpretation criteria for immunohistochemical evaluation of effusion fluids are not modified specifically, it may not be productive in resolving some challenging cases. However, this aspect is not well elaborated in the literature. A basic and critical challenge is finding and locating the cells of interest in cell-block sections of effusion fluids. A unique approach is to choose a fundamental immunopanel which highlight the mesothelial and inflammatory cells in reactive effusion fluids to create the basic map. This allows detection of a ‘second-foreign’ population which can be immunocharacterized further with the help of subtractive coordinate immunoreactivity pattern (SCIP) approach elaborated here.

PD‐L1 immunohistochemical testing: A review with reference to cytology specimens

“…This study did not precisely define the type of specimen undergoing testing for PD‐L1. Formalin fixed paraffin embedded cytologic material may be the optimal sub‐straight for immunohistochemical testing for most antigen targets 22,61,62 . Formalin fixed paraffin embedded cell block material most closely resembles biopsy and resection specimens used for diagnosis and immunohistochemical testing.…”

“…Some authorities believe that alcohol fixation results in impaired immunohistochemical staining 63–66 and should be avoided with primary formalin fixation of cells blocks being the preferred technique. Other authors believe that the primary fixative has little impact of the quality of immunohistochemical staining and alcohol is not deleterious to the validity of PD‐L1 results 9–11,22,55,58–68 . Currently, the optimal technique for preparation of cell blocks remains in question but the majority of studies favor the use of primary formalin fixed paraffin embedded cell block material.…”

Cell-blocks and hematolymphoid lesions

“…e cytomorphology helps in distinguishing endothelial cells (large oval nuclei) that might be CyclinD1 positive from lymphocytes (small, round). Application of SCIP approach [21] and dual color immunostaining [22,23] method using CD20/CyclinD1 may be valuable for proper interpretation. It allows for objective evaluation of this coordinate immunoreactivity for detection of the lymphocyte population with aberrant expression.…”

“…It allows for objective evaluation of this coordinate immunoreactivity for detection of the lymphocyte population with aberrant expression. Both SCIP (Subtractive Coordinate Immunoreactivity Pattern) approach [21] and dual color immunostaining [22,23] methods are relatively simple techniques to be applied by any lab.…”

“…Although criteria for grading of FL by aspiration cytology have been proposed by Young et al; [34] such criteria are not widely applied in clinical practice. Further research on this issue with application of appropriate immunomarkers on qualitatively and quantitatively optimal cell-block [35][36][37] with special approaches including application of SCIP [21] and dual/ multicolor immunostaining [22,23] is recommended.…”