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Product details

Synonyms = Paired box gene 8; Paired domain gene 8; PAX8; paired box homeotic gene 8

Antibody type = Recombinant Rabbit monoclonal / IgG

Clone = MSVA-708R

Positive controlKidney: At least weak to moderate, distinct nuclear staining should be seen in the majority of epithelial cells of the proximal and distal renal tubules, collection ducts, and the parietal epithelial cells of the Bowman’s capsule.

Negative controlTonsil: No staining should be seen in squamous epithelial cells and in lymphocytes.

Cellular localization = Nuclear

Reactivity = Human

 

 

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

Relevance of Antibody

PAX8 is a marker for kidney, thyroid and Mueller tract tissues.

Biology Behind

PAX8 (Paired box gene 8) is a member of the paired-box gene family. The 48kDa protein is coded by the PAX8 gene at chromosome 2q14.1. PAX8 is physiologically expressed in organ development of the thyroid, Wollffian and Muellerian tract, renal/upper urinary tract, and the inner ear, and is also required for tissue homeostasis in the respective adult tissue. PAX8 is a master transcriptional regulator of thyroid specific genes such as thyroglobulin, thyroid peroxidase, and the sodium-iodide symporter by binding to the promoter regions. In Wollffian and Muellerian duct derived tissues, PAX8 is important for mesenchymal-to-epithelial transition and regulates branching morphogenesis and nephron differentiation. PAX8 may also modulate WT1 transcription. Inherited PAX gene deficiencies can result in  congenital hypothyroidism and in genitourinary development defects (Congenital Anomalies of the Kidney and Urinary Tract; CAKUT).

Staining Pattern in Normal Tissues

PAX8 immunostaining preferably occurs in epithelial cells of the kidney, endometrium, endocervix, fallopian tube, and in follicular cells of the thyroid. Images describing the PAX8  staining pattern in normal tissues obtained by the antibody MSVA-708R are shown in our “Normal Tissue Gallery”.

 

Brain Cerebrum Negative.
Cerebellum Negative.
Endocrine Tissues Thyroid Strong nuclear and also a variable cytoplasmic PAX8 staining of follicular cells.
Parathyroid Negative.
Adrenal gland Negative.
Pituitary gland Negative (for nuclear staining). A weak to moderate (non-specific), purely cytoplasmic PAX8 staining can occur in a subset of epithelial cells.
Respiratory system Respiratory epithelium Negative.
Lung Negative.
Gastrointestinal Tract Salivary glands Negative.
Esophagus Negative.
Stomach Negative (for nuclear staining). A faint non-specific and purely cytoplasmic PAX8 staining can occur in stomach glands.
Duodenum Negative. See remarks.
Small intestine Negative. See remarks.
Appendix Negative.
Colon Negative.
Rectum Negative.
Liver Negative.
Gallbladder Negative. A faint non-specific) cytoplasmic staining can occur in epithelial cells.
Pancreas Negative.
Genitourinary Kidney Variable, weak to strong nuclear PAX8 staining of cells of proximal and distal tubuli, collecting ducts, and of epithelial cells of the Bowman’s capsule. Some cytoplasmic staining also occurs in these cells.
Urothelium PAX8 staining is usually absent in urothelium, but a weak to moderate PAX8 positivity can occasionally occur in basal and suprabasal cell layers, especially in the kidney pelvis.
Male genital Prostate Negative.
Seminal vesicles Variable, weak to strong nuclear PAX8 staining of epithelial cells, often accompanied by some cytoplasmic staining.
Testis Negative.
Epididymis Variable, usually strong nuclear PAX8 staining of epithelial cells in the cauda and the caput epididymis.
Female genital Breast Negative.
Uterus, myometrium Negative.
Uterus, ectocervix Negative.
Uterus endocervix Strong nuclear PAX8 staining of epithelial cells.
Uterus, endometrium Variable, usually strong nuclear PAX8 staining of endometrial cells while stroma cells remain negative.
Fallopian Tube Strong nuclear PAX8 staining of a subset of epithelial cells of the fallopian tube
Ovary Negative.
Placenta early Negative.
Placenta mature Negative.
Amnion Negative.
Chorion Negative.
Skin Epidermis Negative.
Sebaceous glands Negative.
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 Nuclear staining obtained by MSVA-708R is completely specific for PAX8. A (non-specific) cytoplasmic staining occurs in a small subset of inflammatory cells, especially in the small intestine as well as few other tissues.

These findings are largely consistent with the RNA data described in the Human Protein Atlas (Tissue expression PAX8).

 

Positive controlKidney: At least weak to moderate, distinct nuclear staining should be seen in the majority of epithelial cells of the proximal and distal renal tubules, collection ducts, and the parietal epithelial cells of the Bowman’s capsule.

Negative controlTonsil: No staining should be seen in squamous epithelial cells and in lymphocytes.

 

Normal tissue gallery

Staining Pattern in Relevant Tumor Types

PAX8 expression primarily occurs in carcinomas derived from the thyroid, kidney, ovary, and the endometrium. At lower frequency, PAX8 expression also occurs in tumors from various other organs of origin.

The TCGA findings on PAX8 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 MSVA-708R 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 role of PAX8 in tumor biology  – especially for epithelial mesenchymal transition – is under investigation.
  • The utility of PAX8 as a therapeutic target is being studied.
  • A possible role of PAX8 as an inducer of angiogenesis is being considered.

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 MSVA-708R specificity for detection of PAX8 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 PAX8). PAX8 positivity by MSVA-708R is detectable in all tissues with documented PAX8 RNA expression (kidney, epididymis, seminal vesicle, endometrium, cervix uteri, fallopian tube, thyroid). All tissues without previously documented PAX8 RNA expression did not show a positive nuclear immunostaining by MSVA-708R

 

Comparison of antibodies: True expression of PAX8 in all cell types for which a nuclear PAX8 positivity was seen by MSVA-708R is corroborated by confirmation of all nuclear PAX8 stainings obtained by MSVA-708R by another commercially available independent antibody (termed “validation antibody”). That the second antibody also showed nuclear staining of lymphocytes and other tissues for which a nuclear staining was not seen by MSVA-706R, documents the independent nature of the validation antibody. These nuclear stainings of the validation antibody are considered antibody specific cross-reactivities. Some purely cytoplasmic stainings seen by MSVA-708R in few inflammatory cells of the intestine, a subset of cells of the adenohypophysis, gallbladder epithelium and gastric glands are also considered cross-reactivities. These, however, cannot cause interpretation issues because they are not nuclear.

Antibody Comparison: MSVA-708R vs another commercial anti-PAX8 antibody called “validation antibody”

 

 

 

 

Normal tissue gallery