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EP1: a novel rabbit monoclonal antibody for detection of oestrogen receptor ?.

ABSTRACT: AIMS: Assessment of hormone receptor expression is part of routine examination of every breast cancer. In this study, we report the characterisation of a novel rabbit monoclonal antibody, clone EP1, directed against oestrogen receptor (ER) ?. Additionally, its immunohistochemical performance characteristics in archival tissues are evaluated in normal tissues and two distinct cohorts of breast cancer patients. METHODS: Comparative analyses between EP1 and the anti-ER? component of the ER/PR pharmDx kit (cocktail of mouse monoclonal antibody clones 1D5 and ER-2-123) and between EP1 and another commercially available rabbit monoclonal antibody, clone SP1, are described. RESULTS: Clone EP1 specifically detects nuclear ER in all tissues examined; cytoplasmic staining was not observed. The analysis shows a high degree of concordance (~95%) between EP1 and both the ER? component of the Dako ER/PR pharmDx kit and Ventana clone SP1. However, the use of EP1 antibody together with Dako EnVision FLEX detection system resulted in a stronger staining intensity as compared with SP1 antibody using the Ventana ultraView DAB detection system resulting in better 'ease of use.' CONCLUSIONS: The use of EPI can result in better interpretation of the results of the ER analysis.

SUBMITTER: Badve S

PROVIDER: S-EPMC3841770 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

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EP1: a novel rabbit monoclonal antibody for detection of oestrogen receptor α.

Badve Sunil S Vladislav I Tudor IT Spaulding Betsy B Strickland Anna A Hernandez Sylvia S Bird-Turner Lisa L Dodson Cecelia C Elleby Bjorn B Phillips Therese T

Journal of clinical pathology 20130726 12

<h4>Aims</h4>Assessment of hormone receptor expression is part of routine examination of every breast cancer. In this study, we report the characterisation of a novel rabbit monoclonal antibody, clone EP1, directed against oestrogen receptor (ER) α. Additionally, its immunohistochemical performance characteristics in archival tissues are evaluated in normal tissues and two distinct cohorts of breast cancer patients.<h4>Methods</h4>Comparative analyses between EP1 and the anti-ERα component of th ...[more]

PMID: 23894168

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Project description:OBJECTIVE:Insulin resistance is a key feature of Type 2 Diabetes (T2D), and improving insulin sensitivity is important for disease management. Allosteric modulation of the insulin receptor (IR) with monoclonal antibodies (mAbs) can enhance insulin sensitivity and restore glycemic control in animal models of T2D. METHODS:A novel human mAb, IRAB-A, was identified by phage screening using competition binding and surface plasmon resonance assays with the IR extracellular domain. Cell based assays demonstrated agonist and sensitizer effects of IRAB-A on IR and Akt phosphorylation, as well as glucose uptake. Lean and diet-induced obese mice were used to characterize single-dose in vivo pharmacological effects of IRAB-A; multiple-dose IRAB-A effects were tested in obese mice. RESULTS:In vitro studies indicate that IRAB-A exhibits sensitizer and agonist properties distinct from insulin on the IR and is translated to downstream signaling and function; IRAB-A bound specifically and allosterically to the IR and stabilized insulin binding. A single dose of IRAB-A given to lean mice rapidly reduced fed blood glucose for approximately 2 weeks, with concomitant reduced insulin levels suggesting improved insulin sensitivity. Phosphorylated IR (pIR) from skeletal muscle and liver were increased by IRAB-A; however, phosphorylated Akt (pAkt) levels were only elevated in skeletal muscle and not liver vs. control; immunochemistry analysis (IHC) confirmed the long-lived persistence of IRAB-A in skeletal muscle and liver. Studies in diet-induced obese (DIO) mice with IRAB-A reduced fed blood glucose and insulinemia yet impaired glucose tolerance and led to protracted insulinemia during a meal challenge. CONCLUSION:Collectively, the data suggest IRAB-A acts allosterically on the insulin receptor acting non-competitively with insulin to both activate the receptor and enhance insulin signaling. While IRAB-A produced a decrease in blood glucose in lean mice, the data in DIO mice indicated an exacerbation of insulin resistance; these data were unexpected and suggested the interplay of complex unknown pharmacology. Taken together, this work suggests that IRAB-A may be an important tool to explore insulin receptor signaling and pharmacology.

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EP1: a novel rabbit monoclonal antibody for detection of oestrogen receptor ?.

Publications

EP1: a novel rabbit monoclonal antibody for detection of oestrogen receptor α.

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