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Product details

Synonyms = glutathione peroxidase 2 , GI-GPx , GPRP , GPRP-2 , GPx-2 , GPx-GI , GSHPX-GI , GSHPx-2

Antibody type = Recombinant Rabbit monoclonal / IgG

Clone = HMV-301

Positive control = Colon: A strong GPX2 staining should be seen in epithelial cells (predominantly in the crypt base).

Negative control = Colon: Stroma cells, inflammatory cells, and smooth muscle cells must be GPX2 negative.

Cellular localization = Nucleus

Reactivity = Human

 

 

Application = Immunohistochemistry
Dilution = 1:100 – 1:200
Intended Use = Research Use Only

Relevance of Antibody

GPX2 is critical for response to oxidative stress.

Biology Behind

The Glutathione peroxidase 2 (GPX2) is a selenium-dependent glutathione peroxidase coded by the GPX2 gene on 14q24.1. GPX2 is one out of eight known glutathione peroxidases (Gpx1-8) in humans. GPX2 has a major role in removing potentially harming reactive oxygen species (ROS) from cells by catalyzing the reduction process of hydrogen peroxide to water. GPX2 expression is restricted to few tissues and it is preferentially expressed in the gut where it makes up for about 50% of the total GPx activity. Studies in mice indicate that GPX2 mRNA levels respond to changes in the luminal microflora, suggesting a role of the ileal glutathione peroxidases in preventing inflammation in the GI tract. Mice that lack both GPX1 and GPX2 develop ileocolitis. GPX2 may also be involved in the control of apoptosis. GPX2 has a yet insufficiently understood role in cancer. Its level of expression was described to be prognostic in several cancer types including bladder, nasopharyngeal and prostate cancer. In hepatocellular carcinomas, GPX2 levels markedly affected the lenvatinib-induced ROS levels and apoptosis in HCC cells and also predicted response to lenvatinib therapy in clinical patients (36725188).

Staining Pattern in Normal Tissues

Images describing the GPX2 staining pattern in normal tissues obtained by the antibody HMV-301 are shown in our “Normal Tissue Gallery”.

Brain Cerebrum Negative.
Cerebellum Negative.
Endocrine Tissues Thyroid Negative.
Parathyroid Negative.
Adrenal gland Negative.
Pituitary gland Negative.
Respiratory system Respiratory epithelium Weak to moderate, nuclear and cytoplasmic GPX2 staining of basal cells.
Lung Negative.
Gastrointestinal Tract Salivary glands Intense, predominantly cytoplasmic GPX2 staining of a variable number of cells of some (but not all) excretory ducts.
Esophagus Weak nuclear and cytoplasmic GPX2 staining of basal and suprabasal squamous epithelial cells.
Stomach GPX2 staining is strong in the surface epithelium and strongest in the most apical (luminal) epithelial cells. Staining is only minimal in gastric glands (exception: patrietal cells).
Duodenum A predominantly cytoplasmic GPX2 staining of epithelial cells predominantes in the crypts. A luminal membrane staining of epithelial cells can also be seen.
Small intestine A predominantly cytoplasmic GPX2 staining of epithelial cells predominantes in the crypts. A luminal membrane staining of epithelial cells can also be seen.
Appendix Strong predominantly cytoplasmic GPX2 staining of epithelial cells – predominantly in the crypts. A luminal membrane staining can also be seen.
Colon Strong predominantly cytoplasmic GPX2 staining of epithelial cells – predominantly in the crypts.
Rectum Strong predominantly cytoplasmic GPX2 staining of epithelial cells – predominantly in the crypts.
Liver Weak nuclear and cytoplasmic GPX2 staining of hepatocytes. Markedly stronger staining of bile ducts.
Gallbladder Intense nuclear and cytoplasmic GPX2 staining of eptheliel cells.
Pancreas Significant, predominantly cytoplasmic GPX2 staining of excretory duct cells. Weak, predominantly nuclear staining of pancreatic islet cells.
Genitourinary Kidney Negative.
Urothelium Intense nuclear and cytoplasmic GPX2 staining of urothelial cells. Staining is markedly less intense and may even be absent in umbrella cells.
Male genital Prostate Weak to moderate, nuclear and cytoplasmic GPX2 staining of a subset of basal cells.
Seminal vesicles Moderate to strong, nuclear and cytoplasmic GPX2 staining of a subset of basal cells.
Testis Negative.
Epididymis Weak to moderate, nuclear and cytoplasmic GPX2 staining of basal cells in the corpus.
Female genital Breast Negative.
Uterus, myometrium Negative.
Uterus, ectocervix A variable, weak to moderate nuclear and cytoplasmic GPX2 staining occurs in basal and suprabasal cells.
Uterus endocervix Negative.
Uterus, endometrium Negative.
Fallopian Tube Negative.
Ovary Negative.
Placenta early Negative.
Placenta mature Negative.
Amnion Negative.
Chorion Negative.
Skin Epidermis A variable nuclear and cytoplasmic GPX2 staining can be seen in the epidermis. If present, the staining intensity may decrease from basal to apical cell layers.
Sebaceous glands A weak to moderate nuclear and cytoplasmic staining of peripheral germinative cells of sebaceous glands can be seen.
Muscle/connective tissue Heart muscle Negative.
Skeletal muscle Negative.
Smooth muscle Negative.
Vessel walls Negative.
Fat Negative.
Stroma Negative.
Endothelium Negative.
Bone marrow/ lymphoid tissue Bone marrow Negative.
Lymph node Negative.
Spleen Negative.
Thymus Negative.
Tonsil Negative.
Remarks

Cyclin E1 staining was always nuclear. A particularly strong Cyclin E1 positivity in a large fraction of cells occurred in chorion cells, cytotrophoblast cells, and decidua cells of the placenta, epithelial cells of proliferating endometrium, superficial cell layers of the urothelium and squamous epithelium (especially skin and tonsil). A weak to moderate staining was also seen in epithelial cells of the adrenal cortex and of the parathyroid gland. A faint to weak to moderate Cyclin E1 staining could further be observed in virtually every tissue in cell types/tissue areas with proliferative activity. These for example included tissues with inflammation or reparation, crypt cells in the intestine, neck cells of the stomach mucosa, lymphocytic cells in germinal centres or the thymus, spermatocytes of the testis, amnion cells, and hematopetic cells of the bone marrow. A particular strong Cyclin E1 occurred in chorion cells, cytotrophoblast cells, and decidua cells of the placenta, epithelial cells of proliferating endometrium, superficial cell layers of the urothelium and squamous epithelium (especially skin and tonsil). Tissues with particularly low or absent Cyclin E1 expression included, skeletal and heart muscle, the brain, pituitary gland, thyroid, prostate, seminal vesicle, epididymis, kidney, breast, endocervix, fallopian tube, lung, respiratory epithelium.

The findings described above are thus consistent with the RNA data described in the Human Protein Atlas (Tissue expression GPX2). GPX2 staining by HMV-301 is most prominent in tissues from the gastrointestinal tract but also occurs – at lower levels – in various other tissues.

 

Positive controlColon: A strong GPX2 staining should be seen in epithelial cells (predominantly in the crypt base).

Negative control = Colon: Stroma cells, inflammatory cells, and smooth muscle cells must be GPX2 negative.

 

Normal tissue gallery

Staining Pattern in Relevant Tumor Types

GPX2 is primarily expressed in carcinomas of the gastrointestinal tract, urothelial neoplasms and in squamous cell carcinomas of various organs of origin. GPX2 expression may also occur in other tumors.

The TCGA findings on GPX2 RNA expression in different tumor categories have been summarized in the Human Protein Atlas.

 

 

Cancer tissue gallery

Compatibility of Antibodies

No data available at the moment

Protocol Recommendations

IHC users have different preferences on how the stains should look like. Some prefer high staining intensity of the target stain and even accept some background. Others favor absolute specificity and lighter target stains. Factors that invariably lead to more intense staining include higher concentration of the antibody and visualization tools, longer incubation time, higher temperature during incubation, higher temperature and longer duration of the heat induced epitope retrieval (slide pretreatment). The impact of the pH during slide pretreatment has variable effects and depends on the antibody and the target protein.

 

All images and data shown here and in our image galleries are obtained by the manual protocol described below. Other protocols resulting in equivalent staining are described as well.

 

Manual protocol

Freshly cut sections should be used (less than 10 days between cutting and staining). Heat-induced antigen retrieval for 5 minutes in an autoclave at 121°C in pH 7,8 Target Retrieval Solution buffer. Apply HMV-301 at a dilution of 1:150 at 37°C for 60 minutes. Visualization of bound antibody by the EnVision Kit (Dako, Agilent) according to the manufacturer’s directions.

Potential Research Applications

  • The diagnostic and prognostic relevance of GPX2 expression in tumors and in preneoplastic disease needs to be investigated.
  • GPX2 may serve as a therapeutic target. 

Evidence for Antibody Specificity in IHC

There are two ways how the specificity of antibodies can be documented for immunohistochemistry on formalin fixed tissues. These are: 1. Comparison with a second independent method for target expression measurement across a large number of different tissue types (orthogonal strategy), and 2. Comparison with one or several independent antibodies for the same target and showing that all positive staining results are also seen with other antibodies for the same target (independent antibody strategy). 

 

Orthogonal validation: For the antibody HMV-301 specificity is suggested by the strong concordance of the immunostaining data with data from three independent RNA screening studies, including the Human Protein Atlas (HPA) RNA-seq tissue dataset, the FANTOM5 project, and the Genotype-Tissue Expression (GTEx) project, which are all summarized in the Human Protein Atlas (Tissue expression GPX2)

 

GPX2 positivity by HMV-301 is strongest in the tissues with highest documented GPX2 RNA expression (stomach, duodenum, small intestine, appendix, colon, rectum, gallbladder, urinary bladder). That GPX2 staining is less intense in the liver where high RNA expression levels are also seen is explained by the fact that – in contrast to the intestine – nearly all liver cells (hepatocytes) express some GPX2 which may cause a high average level of GPX2 RNA in liver tissue. GPX2 positivity by HMV-301 is also detectable in the tissues with lower level GPX2 RNA expression (skin, ectocervix, esophagus, salivary glands, pancreas, prostate, seminal vesicles). Moreover, GPX2 staining by HMV-301 is lacking in most tissues not showing RNA expression (spleen, lymph node, bone marrow, thymus, tonsil, skeletal muscle, heart muscle, placenta, breast, fallopian tube, ovary, endometrium, testis, endocrine organs, brain). The few tissues without previously documented GPX2 RNA expression but GPX2 positivity by HMV-301 had either very few positive cells that were probably not detected in RNA studies (basal cells of the cauda epididymis) or were previously not analyzed on the RNA level (respiratory epithelium).

 

Comparison of antibodies: True expression of GPX2 in all cell types found GPX2 positive by HMV-301 (including basal cells of the cauda epididymis and respiratory epithelium) is corroborated by identical stainings obtained by another commercially available independent antibody (termed “validation antibody”). The additional staining of nuclei of various tissues (for example: pneumocytes, chorion cells and trophoblastic cells of the placenta, pituicytes of the neurohypophysis) by the “validation antibody” demonstrates the independence of the validation antibody. 



 

 

 

 

 

 

Normal tissue gallery