Abstract
Pre-analytical factors, such as fixation time, influence morphology of diagnostic and predictive immunohistochemical staining, which are increasingly used in the evaluation of lung cancer. Our aim was to investigate if variations in fixation time influence the outcome of immunohistochemical staining in lung cancer. From lung resections, specimen with tumor size bigger than 4 cm, 10 samples were obtained: 2 were put through the standard fixation protocol, 5 through the delayed, and 3 through the prolonged fixation protocol. After paraffin embedding, tissue microarrays (TMAs) were made. They were stained with 20 antibodies and scored for quality and intensity of staining. Samples with delay in fixation showed loss of TMA cores on glass slides and deterioration of tissue quality leading to reduction in the expression of CK 7, Keratin MNF116, CAM 5.2, CK 5/6, TTF-1, C-MET, Napsin A, D2-40, and PD-L1. Prolonged fixation had no influence on the performance of immunohistochemical stains. Delay of fixation negatively affects the expression of different immunohistochemical markers, influencing diagnostic (cytokeratins) and predictive (PD-L1) testing. These results emphasize the need for adequate fixation of resection specimen.
Highlights
Tissue samples are used for diagnostic and predictive analysis
There is a long list of variables in tissue handling, highly varying around the world [1], from the moment tissue is removed from a patient to formalin fixation and from formalin fixation to paraffin embedded (FFPE) tissue [2], influencing tissue quality
“Time before fixation,” as a very important pre-analytical factor, influences degradation of proteins and may cause reduced binding of the primary antibody. It has already been shown in breast cancer that delay in fixation influences the outcome of immunohistochemical staining [7, 8]
Summary
Tissue samples are used for diagnostic and predictive analysis. Fixation process, preventing degradation, is essential in preserving the tissue. In the last 3–4 decades, immunohistochemistry (IHC) has become an essential procedure in pathology diagnostics [3,4,5] It is important for tumor classification and characterization of underlying agents in many infectious diseases [6] and, in addition, is increasingly used in predictive testing [4]. “Time before fixation,” as a very important pre-analytical factor, influences degradation of proteins and may cause reduced binding of the primary antibody. It has already been shown in breast cancer that delay in fixation influences the outcome of immunohistochemical staining [7, 8]
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