Development of Ki-67 rapid immunocytochemistry and its evaluation in intraoperative squash smear diagnosis of central nervous system gliomas: Study protocol for a prospective and observational study

An intraoperative diagnosis of central nervous system (CNS) gliomas, particularly differentiating low-grade from high-grade glioma, plays a crucial role in guiding the surgeon’s decisions per operatively. Immediate microscopic examination of fresh tumor tissue is commonly done by squash smear cytology preparation, which is a rapid, simple, and technically easy procedure for studying the cellular morphology. However, accurate grading of a glioma, particularly CNS World Health Organization grade 2 and grade 3 tumors which often have overlapping features, is a common diagnostic difficulty confronted in the limited time and tissue material available during an intraoperative procedure. “Development of Ki-67 Rapid Immunocytochemistry and its Evaluation in Intraoperative Squash Smear Diagnosis of CNS Gliomas” is a prospective and observational study, where we aim to develop a protocol for performing rapid immunocytochemistry (RICC) for Ki-67/MIB-1 proliferation index on intraoperative squash smears and compare its efficacy with conventional immunohistochemistry (IHC) in aiding diagnosis of CNS gliomas. Patients clinically suspected with CNS glioma will be undergoing intra-operative squash smear analysis. Smears for cytological evaluation shall be made from the tissue obtained for intra-operative diagnosis and stained with rapid hematoxylin and eosin stain for a morphological diagnosis. An additional unstained smear shall next be subjected to RICC for the Ki-67 proliferation index. A novel cost-effective in-house validated protocol for RICC shall be developed to deliver the Ki-67 proliferation index. The primary outcome shall be the agreement between Ki-67 proliferation indices obtained by RICC on squash smears and conventional IHC performed on formalin-fixed paraffin-embedded sections of the tumor tissue for its diagnostic accuracy.