MUC1 CLIA Kit

Catalog No. ABIN504789

Product Details MUC1 CLIA Kit

Target: MUC1 (Mucin 1 (MUC1))

Reactivity: Human

Detection Method: Chemiluminescent

Method Type: Sandwich ELISA

Application: ELISA

Purpose: Immunoenzymometric sequential assay (TYPE 4): The essential reagents required for an immunoenzymometric assay include high affinity and specificity antibodies (enzyme and immobilized), with different and distinct epitope recognition, in excess, and native antigen. In this procedure, the immobilization takes place during the assay at the surface of a microplate well through the interaction of streptavidin coated on the well and exogenously added biotinylated monoclonal anti-CA15-3 antibody. Upon mixing monoclonal biotinylated antibody, and a serum containing the native antigen, reaction results between the native antigen and the antibody, forming an antibody-antigen complex.

Analytical Method: Quantitative

Characteristics: The Quantitative Determination of Cancer Antigen (CA 15-3) Concentration in Human Serum by a Microplate Chemiluminescence assay

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. For accurate comparison to establish normal values, a fasting morning serum sample should be obtained. 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 diluted specimen is required.

Reagent Preparation:

1. Wash Buffer Dilute contents of wash solution to 1000ml with distilled or deionized water in a suitable storage container. Diluted buffer can be stored 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. 3. Patient Sample Dilution (1:21) Dispense 25 (l of each control and/or patient specimen into 0.500 ml of CA 15-3 dilution matrix appropriately labeled, clean container(s) and mix thoroughly before use. Store refrigerated at 2-8(C for up to 48 hours.

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 diluted serum reference, control or specimen into the assigned well. 3. Add 0.100 ml (100l) of the biotinylated labeled antibody to each well. It is very important to dispense all reagents close to the bottom of the coated well. 4. Swirl the microplate gently for 20-30 seconds to mix and cover. 5. Incubate 45 minutes at room temperature. 6. Discard the contents of the microplate by decantation or aspiration. If decanting, tap and blot the plate dry with absorbent paper. 7. Add 350l of wash buffer (see Reagent Preparation Section), decant (tap and blot) or aspirate. Repeat two (2) additional times for a total of three (3) 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 two (2) additional times. 8. Add 0.100 ml (100l) of the Ca15-3 Tracer Reagent to each well. DO NOT SHAKE THE PLATE AFTER TRACER ADDITION 9. Cover and incubate 45 minutes at room temperature. 10. Discard the contents of the microplate by decantation or aspiration. If decanting, blot the plate dry with absorbent paper. 11. Add 350l of wash buffer (see Reagent Preparation Section), decant (tap and blot) or aspirate. Repeat two (2) additional times for a total of three (3) 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 two (2) additional times. 12. Add 0.100 ml (100l) of working signal reagent to all wells (see Reagent Preparation Section). ). Always add reagents in the same order to minimize reaction time differences between wells. 13. Incubate for five (5) minutes in the dark. 14. Read the relative light units in each well for 0.2 1.0 seconds. The results should be read within thirty (30) minutes of adding the substrate solution.

Restrictions: For Research Use only

Handling

Target Details

Target: MUC1 (Mucin 1 (MUC1))

Alternative Name: CA 15-3 (MUC1 Products)

Synonyms: CA 15-3 CLIA Kit, CD227 CLIA Kit, EMA CLIA Kit, H23AG CLIA Kit, KL-6 CLIA Kit, MAM6 CLIA Kit, MCKD1 CLIA Kit, MUC-1 CLIA Kit, MUC-1/SEC CLIA Kit, MUC-1/X CLIA Kit, MUC1/ZD CLIA Kit, PEM CLIA Kit, PEMT CLIA Kit, PUM CLIA Kit, Muc-1 CLIA Kit, mucin 1, cell surface associated CLIA Kit, mucin 1, transmembrane CLIA Kit, MUC1 CLIA Kit, Muc1 CLIA Kit

Background: Although multiple serum based tumor markers have been described for breast cancer, such as CA 15-3, BR 27-29, carcinoembryonic antigen (CEA), tissue polypeptide antigen (TPA), tissue polypeptide specific antigen, and HER-2 (the extracellular domain), the most widely used are CA 15-3 and CEA. CA 15-3 is considered to be one of the first circulating prognostic factors for breast cancer (1). Preoperative concentrations thus might be combined with prognostic factors for predicting outcome in patients with newly diagnosed breast cancer(2). At present the most important clinical application of CA 15-3 is in monitoring therapy in patients with advanced breast cancer that is not accessible by existing clinical or radiologic procedures(3). The CA 15-3 assay measures the protein product of MUC1 gene. MUC1 protein is a large transmembrane glycosylated molecule containing three main domains, a large extracellular region, a membrane spanning sequence, and a cytoplasmic domain (4). Although the physiologic function of MUC1 is unclear, the glycoprotein has been implicated in cell adhesion, immunity and metastasis. Compared with healthy breast tissue, MUC1 is present in higher concentrations but less glycosylated in breast carcinoma (5-8). In this method, a prediluted CA15-3 calibrator diluted patient specimen or control is first added to a streptavidin coated well. Biotinylated monoclonal antibody (specific for CA15-3) is added and the reactants mixed. Reaction between the CA15-3 antibodies and native CA15-3 forms complex that binds with the streptavidin coated to the well. The excess serum proteins are washed away via a wash step. Another enzyme labeled antibody specific for a different epitopic recognition of CA15-3 is added to the wells. The enzyme labeled antibody binds to the CA15-3 already immobilized on the well through its binding with the biotinylated monoclonal antibody. Excess enzyme is washed off via a wash step. Light is generated by the addition of a substrate. The intensity of the light generation is directly proportional to the concentration of the CA15-3 in the sample.

Pathways: Negative Regulation of intrinsic apoptotic Signaling