2-Methoxyestradiol ELISA kit

Sample

plasma, urine, other sample matrices

Intended Use

This 2-Methoxyestradiol EIA Kit is a competitive assay that can be used for quantification of 2-ME2 in plasma, urine, and other sample matrices. The EIA typically displays an IC50 (50% B/B0) of approximately 350 pg/mL and a detection limit (80% B/B0) of approximately 40 pg/mL.

Contents of Kit

Storage

For more detailed information, please download the following document on our website.

Performance Characteristics

Sample Data
The standard curve presented here is an example of the data typically obtained with this kit; however, your results will not be identical to these. You must run a new standard curve. Do not use the data below to determine the value of your samples. Your results could differ substantially.



Precision:
The intra- and inter-assay CVs have been determined at multiple points on the standardcurve. These data are summarized in the graph on page 28 and in the table below.

General Description

2-Methoxyestradiol (2-ME2) is a natural metabolite of estradiol with potent antitumor and antiangiogenic properties. Several clinical trials have demonstrated the antitumor effects of 2-ME2 in prostate cancer, breast cancer, renal cell carcinoma, ovarian cancer, glioblastoma, and multiple myeloma. Other studies indicate that 2-ME2 exhibits more potent cardioprotective effects than estradiol. 2-ME2 is produced by cytochrome P450-dependant hydroxylation of 17b-estradiol at the 2-position, followed by O-methylation catalyzed by catechol-O-methyltransferase. Deficiency in catechol-O-methyltransferase and 2-ME2 is associated with pre-eclampsia. Therefore, 2-ME2 may be utilized as a plasma and urinary diagnostic marker for this disease, as well as a therapeutic supplement for its prevention. 2-ME2 has very low affinity for estrogen receptors compared to estradiol and other estradiol metabolites, suggesting a different mechanism of action for this compound. Levels of 2-ME2 in plasma vary from less than 0.1 ng/mL in males and non-pregnant females to 3.0 ng/mL in pregnant women. Urinary excretion of 2-ME2 in non-pregnant women was reported to be 0.91-2.42 nanomoles per 24 hours, which increases dramatically during pregnancy.
This assay is based on the competition between free 2-ME2 and a 2-Methoxyestradiol Tracer (2-ME2 linked to an acetylcholinesterase (AChE) molecule) for a limited number of 2-ME2- specific rabbit antiserum binding sites. The concentration of the 2-Methoxyestradiol Tracer is held constant while the concentration of free 2-ME2 (standard or sample) varies. Thus, the amount of 2-Methoxyestradiol Tracer that is able to bind to the rabbit antiserum will be inversely proportional to the concentration of free 2-ME2 in the well. This rabbit antiserum- 2-ME2 (either free or tracer) complex binds to the mouse monoclonal anti-rabbit IgG that has been previously attached to the well. The plate is washed to remove any unbound reagents and then Ellman's Reagent (which contains the substrate to AChE) is added to the well. The product of this enzymatic reaction has a distinct yellow color and absorbs strongly at 412 nm. The intensity of this color, determined spectrophotometrically, is proportional to the amount of 2-Methoxyestradiol Tracer bound to the well, which is inversely proportional to the amount of free 2-ME2 present in the well during the incubation; or
Absorbance ∝ [Bound 2-Methoxyestradiol Tracer] ∝ 1/[2-ME2]
A schematic of this process is shown below in Figure 1.