Differential detection of cytoplasmic Wilms tumor 1 expression by immunohistochemistry, western blotting and mRNA quantification

Abstract

Abstract. Wilms tumor 1 (WT1) is considered to be a promising target of cancer treatment because it has been reported to be frequently expressed at high levels in various malignancies. Although WT1-targeted cancer treatment has been initiated, conclusive detection methods for WT1 are not established. The present study aimed to consolidate immunohistochemistry for WT1 with statistical basis. Transfected cells with forced WT1 expression yielded specific western blot bands and nuclear immunostaining; cytoplasmic immunostaining was not specifically recognized. Immunohistochemistry, western blotting, and quantitative reverse transcriptase-polymerase chain reaction were performed in 35 human cell lines using multiple WT1 antibodies and their results were quantified. Relationships among the quantified results were statistically analyzed; the nuclear immunostaining positively correlated with western blot bands and mRNA expression levels, whereas cytoplasmic immunostaining did not. These results indicate that nuclear immunostaining reflects WT1 expression but cytoplasmic immunostaining does not. The nuclear immunostaining was barely (3/541) observed in primary cancer of esophagus, bile duct, pancreas and lung. Although the present study has some limitations, the results indicate that the cytoplasmic immunostaining does not correlate with actual WT1 expression and prompts researchers to carefully evaluate target molecule expression in treatment of cancer.

The cell tube block technique and an immunohistochemistry panel including Wilms tumor 1 to assist in diagnosing cavitary effusions in dogs and cats

“…Unlike other tumor suppressor genes, WT1 appears to be expressed in selected mesoderm‐derived cell types, such as the mesothelium, kidney, and gonads . In humans, WT1 is considered a useful mesothelial marker, with positive rates of 100% for reactive mesothelium and between 72% and 100% of mesotheliomas . In our CTB case series and the validation study, WT1 stained positively for all cases of normal, reactive mesothelium, and mesotheliomas (including cases negative for vimentin).…”

“…Several monoclonal and polyclonal antibodies against WT1 are available with different recognition sites along the amino acid sequence . Recently, it was reported that monoclonal antibodies (and particularly the clone used herein) that stain the entire nucleus correlated with Western blot band intensities and mRNA expression levels . In contrast, no correlation was observed with cytoplasmic immunostaining (regardless of which antibody was used) and was considered as nonspecific finding .…”

“…WT1 is a tumor suppressor gene, located on chromosome 18 in the dog . Unlike other tumor suppressor genes, WT1 appears to be expressed in selected mesoderm‐derived cell types, such as the mesothelium, kidney, and gonads . In humans, WT1 is considered a useful mesothelial marker, with positive rates of 100% for reactive mesothelium and between 72% and 100% of mesotheliomas .…”

“…Wilms tumor 1 protein has a typical nuclear staining pattern in formalin‐fixed materials and stains, in most cases, more than 50% of cells similar to what was seen in this study. For WT1, the type of antibody used and expression patterns are particularly relevant, and these may account for some controversy regarding WT1 IHC in different tissues and tumors . Several monoclonal and polyclonal antibodies against WT1 are available with different recognition sites along the amino acid sequence .…”

“…For WT1, the type of antibody used and expression patterns are particularly relevant, and these may account for some controversy regarding WT1 IHC in different tissues and tumors . Several monoclonal and polyclonal antibodies against WT1 are available with different recognition sites along the amino acid sequence . Recently, it was reported that monoclonal antibodies (and particularly the clone used herein) that stain the entire nucleus correlated with Western blot band intensities and mRNA expression levels .…”