Estradiol (17-Estradiol) CLIA Kit

Catalog No. ABIN504761

Product Details

Target: Estradiol (17-Estradiol)

Reactivity: Human

Detection Method: Chemiluminescent

Method Type: Competition ELISA

Application: ELISA

Purpose: Delayed Competitive Enzyme Immunoassay (TYPE 9): The essential reagents required for a enzyme immunoassay include antibody, enzyme-antigen conjugate and native antigen. Upon mixing the biotinylated antibody with a serum containing the antigen, a reaction results between the antigen and the antibody.

Analytical Method: Quantitative

Characteristics: The Quantitative Determination of Estradiol Concentration in Human Serum or Plasma by a Microplate Chemiluminescence Immunoassay

Application Details

Application Notes: All products that contain human serum have been found to be non-reactive for Hepatitis B Surface Antigen, HIV 1&2 and HCV Antibodies by FDA required tests. Since no known test can offer complete assurance that infectious agents are absent, all human serum products should be handled as potentially hazardous and capable of transmitting disease. Good laboratory procedures for handling blood products can be found in the Center for Disease Control / National Institute of Health, "Biosafety in Microbiological and Biomedical Laboratories," 2nd Edition, 1988, HHS Publication No. (CDC) 88-8395.

Plate: Pre-coated

Protocol:

Specimien Collection and Preparation:

The specimens shall be blood, serum or heparanised plasma in type and taken with the usual precautions in the collection of venipuncture samples. The blood should be collected in a redtop (with or without gel additives) venipuncture tube or for plasma use evacuated tube(s) containing heparin. Allow the blood to clot for serum samples. Centrifuge the specimen to separate the serum or plasma from the cells. Samples may be refrigerated at 2_x001E_8oC for a maximum period of five (5) days. If the specimen(s) cannot be assayed within this time, the sample(s) may be stored at temperatures of _x001E_20oC for up to 30 days. Avoid repetitive freezing and thawing. When assayed in duplicate, 0.050ml of the specimen is required.

Reagent Preparation:

1. Wash Buffer Dilute contents of Wash Concentrate to 1000ml with distilled or deionized water in a suitable storage container. Store diluted buffer at room temperature 20-27(C for up to 60 days. 2. Working Signal Reagent Solution - Store at 2 - 8(C. Determine the amount of reagent needed and prepare by mixing equal portions of Signal Reagent A and Signal Reagent B in a clean container. For example, add 1 ml of A and 1ml of B per two (2) eight well strips (A slight excess of solution is made). Discard the unused portion if not used within 36 hours after mixing. If complete utilization of the reagents is anticipated, within the above time constraint, pour the contents of Signal Reagent B into Signal Reagent A and label accordingly.

Test Procedure:

Before proceeding with the assay, bring all reagents, serum references and controls to room temperature (20 - 27(C). 1. Format the microplates wells for each serum reference, control and patient specimen to be assayed in duplicate. Replace any unused microwell strips back into the aluminum bag, seal and store at 2-8(C. 2. Pipette 0.025 ml (25 L) of the appropriate serum reference, control or specimen into the assigned well. 3. Add 0.050 ml (50l) of the Estradiol Biotin Reagent to all wells. 4. Swirl the microplate gently for 20-30 seconds to mix. 5. Cover and incubate for 30 minutes at room temperature. 6. Add 0.050 ml (50l) of Estradiol Tracer Reagent to all wells. Add directly on top the reagents dispensed in the wells 7. Swirl the microplate gently for 20-30 seconds to mix. 8. Cover and incubate for 60 minutes at room temperature. 9. Discard the contents of the microplate by decantation or aspiration. If decanting, blot the plate dry with absorbent paper. 10 Add 350l of wash buffer (see Reagent Preparation Section), decant (tap and blot) or aspirate. Repeat four (4) additional times for a total of five (5) washes. An automatic or manual plate washer can be used. Follow the manufacturers instruction for proper usage. If a squeeze bottle is employed, fill each well by depressing the container (avoiding air bubbles) to dispense the wash. Decant the wash and repeat four (4) additional times. 11. Add 0.100 ml (100l) of working signal reagent solution to all wells (see Reagent Preparation Section). Always add reagents in the same order to minimize reaction time differences between wells. 12. Incubate at room temperature for five (5) minutes in the dark. 13. Read the relative light units in each well with a chemiluminescence microplate reader for 0.5-1.0 seconds. The results should be read within 30 minutes after adding the working Signal Reagent. Note: Dilute the samples suspected of concentrations higher than 3000pg/ml 1:5 and 1:10 with estradiol 0 pg/ml calibrator or male patient serum pools with a known low value for estradiol.

Restrictions: For Research Use only

Handling

Target Details

Target: Estradiol (17-Estradiol)

Background: Measurement of estradiol in serum or plasma is considered to be the most reliable way to assess its rate of production. Estradiol (17-estradiol) is a steroid hormone (molecular weight of 272.3 daltons), which circulates predominantly protein-bound. In addition to estradiol, other natural steroidal estrogens include estrone, estriol and their metabolites. Natural estrogens are hormones secreted principally by the ovarian follicles and also by the adrenals, corpus luteum, and placenta and, in males, by the testes. Exogenous estrogens (natural or synthetic) elicit, to varying degrees, all the pharmacologic responses usually produced by endogenous estrogens. Estrogenic hormones are secreted at varying rates during the menstrual cycle throughout the period of ovarian activity. During pregnancy, the placenta becomes the main source of estrogens. At menopause, ovarian secretion of estrogens declines at varying rates. The gonadotropins of the anterior pituitary regulate secretion of the ovarian hormones, estradiol and progesterone, hypothalamic control of pituitary gonadotropin production is in turn regulated by plasma concentrations of the estrogens and progesterone. This complex feedback system results in the cyclic phenomenon of ovulation and menstruation. Estradiol determinations have proved of value in a variety of contexts, including the investigation of precocious puberty in girls and gynecomastia in men. Its principal uses have been in the differential diagnosis of amenorrhea and in the monitoring of ovulation induction. This kit uses a specific anti-estradiol antibody, and does not require prior sample extraction of serum or plasma. Cross-reactivity to other naturally occurring and structurally related steroids is low. The employment of several serum references of known estradiol concentration permits construction of a graph of activity(light) and con_x001F_centration. From comparison to the dose response curve, an unknown specimen's activity can be correlated with estradiol concentration.