Comparison of Fixation Methods for Preservation of Morphology, RNAs, and Proteins From Paraffin-Embedded Human Cancer Cell-Implanted Mouse Models

Abstract

SummaryXenograft transplantation of human tumor cells into immunodeficient mice is an important method to clarify the roles of specific molecules or chemicals in vivo. Recently, this method has been reported as a definitive examination to identify tumor stem cells. In this study, the authors compared the morphology and the quality and quantity of ribonucleic acid (RNA) and protein in paraffin-embedded tissues of nude mice implanted with human uterine cervical cancer cells, followed by fixation with commonly used fixatives, including 4% paraformaldehyde (PFA), 10% neutral buffered formalin (NBF), 20% NBF, and 99% ethanol (EtOH). The quality of the isolated RNA from PFA-and NBF-fixed paraffin-embedded tissues was high, while EtOH-fixed tissues showed degradation of RNA. NBF-fixed tissues showed excellent quality of morphology, but EtOH-fixed tissues showed contraction of cells. Immunohistochemical results showed differences depending on fixations. The 99% EtOH-fixed samples showed decreases of Ki-67 and VEGF-A immunoreactivities, but improved cytokeratin immunoreactivity. This study indicated that formalin fixation is better than alcohol fixation for RNA preservation in paraffinembedded cancer cell implantation models. Immunohistochemical results differed markedly depending on fixation materials and antibodies; therefore, suitable fixations are needed to quantify and compare the results of immunohistochemical staining on cancer cell implanted nude mice tissues. (J Histochem Cytochem 59:68-75, 2011)

Enhanced detection of Porcine reproductive and respiratory syndrome virus in fixed tissues by in situ hybridization following tyramide signal amplification

“…9,19 In the present study, PRRSV was not detected by IHC in tissue samples fixed for more than 14 days in either 4% PFA or 10% NBF. Prolonged formalin fixation has been shown to lead to masking of epitopes because of cross-linking, which reduces or even abolishes antigen detection.…”

“…[6][7][8]21 Formalin is a commonly used fixative agent that inactivates most infectious agents and inhibits autolysis. 9,19 Formalin fixation results in the cross-linking of tissue protein with DNA or RNA; overfixation can reduce the sensitivity of IHC and ISH. For routine IHC and ISH, solutions of 10% neutral buffered formalin (NBF) or 4% paraformaldehyde (PFA) are commonly used for fixation.…”

Nestin as a novel therapeutic target for pancreatic cancer via tumor angiogenesis

“…Eight weeks later, 2 mice exhibited abdominal tumors in the area adjacent to the pancreas, and the pancreatic tumors were excised. Portions of the subcutaneous and pancreatic tumors were fixed in 4% paraformaldehyde for routine histopathological processing (40). Animal experiments were carried out according to the institutional animal care guidelines of the Nippon Medical School Animal Ethics Committee.…”

An Infant With Non-Ketotic Hyperglycemia: A Case Report

“…Tissue specimens can be kept for a long time after fixing in formalin and embedding in paraffin (FFPE). Nowadays, formaldehyde is used as a common chemical tissue fixator, which is widely used due to low cost and minimal changes in cell and tissue structure (4,5). To preserve the tissue structure during fixation in formalin, the tissues are embedded in paraffin.…”