beta-2 Microglobulin ELISA Kit

Catalog No. ABIN956252

Product Details beta-2 Microglobulin ELISA Kit

Target: beta-2 Microglobulin (B2M) (beta-2-Microglobulin (B2M))

Reactivity: Rat

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Application: ELISA

Purpose: The Rat Beta 2-Microglobulin ELISA is a highly sensitive two-site enzyme-linked immunoassay (ELISA) for the quantitative determination of Beta 2-Microglobulin in rat biological fluids.

Analytical Method: Quantitative

Characteristics: In this assay the B2M present in the sample reacts with the anti-B2M antibody which has been adsorbed to the surface of polystyrene microtiter wells. After the removal of unbound sample proteins by washing, anti-B2M antibody conjugated with horseradish peroxidase (HRP) is added. This HRP-conjugated antibody forms a complex with the previously bound B2M. Following another washing step, the enzyme bound to the immunosorbent is assayed by the addition of a chromogenic substrate, 3,3',5,5'- tetramethylbenzidine (TMB). The quantity of bound enzyme is proportional to the concentration of B2M in the sample tested, thus, the absorbance, at 450 nm, is a measure of the concentration of B2M in the test sample. The quantity of B2M in the test sample can be interpolated from the calibration curve constructed from the calibrators, and corrected for sample dilution.

Components: 1. Diluent Concentrate: One bottle containing 50 mL of a 5X concentrated diluent running buffer.
2. Wash Solution Concentrate: One bottle containing 50 mL of a 20X concentrated wash solution.
3. Enzyme-Antibody Conjugate Concentrate: One vial containing 150 µL of a 100X concentrated affinity-purified anti-rat B2M antibody conjugated with HRP in a stabilizing buffer.
4. TMB Substrate Solution: One vial containing 12 mL of TMB and hydrogen peroxide in citric acid buffer at pH 3.3.
5. Stop Solution: One vial containing 12 mL of 0.3 M sulfuric acid. WARNING: Avoid contact with skin.
6. Microtiter Plate: Twelve removable eight-well strips in well holder frame. Wells are coated with affinity-purified anti-rat B2M.
7. Rat B2M Calibrator: One vial containing a Rat B2M Calibrator.
8. Positive Control: One vial containing 50 µL of serum with 0.1% sodium azide. See the Control Certificate for the concentration.

Material not included: Precision pipettes (2 µL to 200 µL) for making and dispensing dilutions
Test tubes
Microplate washer/aspirator
Distilled or de-ionized H2O Microplate reader
Assorted glassware for the preparation of reagents and buffer solutions
Timer
Vortex mixer

Application Details

Plate: Pre-coated

Reagent Preparation:

1. Diluent Concentrate: The Diluent solution supplied is a 5X concentrate and must be diluted 1:5 with distilled or de-ionized water.
   2. Wash Solution Concentrate: The Wash Solution supplied is a 20X concentrate and must be diluted 1:20 with distilled or de-ionized water. Crystal formation in the concentrate is not uncommon when storage temperatures are low. Warming of the concentrate to 30-35°C before dilution can dissolve crystals.
   3. Enzyme-Antibody Conjugate Concentrate: Calculate the required amount of working conjugate solution for each microtiter plate test strip by adding 10 µL Enzyme-Antibody Conjugate to 990 µL of 1X Diluent for each test strip to be used for testing. Mix uniformly, but gently. Avoid foaming.
   4. TMB Substrate Solution: Ready to use as supplied.
   5. Stop Solution: Ready to use as supplied.
   6. Microtiter Plate: Ready to use as supplied. Unseal Microtiter Pouch and remove plate from pouch. Remove all strips and wells that will not be used in the assay and place back in pouch with desiccant and re-seal.
   7. Rat B2M Calibrator: The Rat B2M Calibrator should be aliquoted and stored frozen. It is at a concentration of 1.60 µg/mL and needs to be diluted in 1X diluent according to the chart below for each run. Rat B2M calibrators need to be prepared immediately prior to use. Mix well between each step. Avoid foaming.
   8. Positive Control: The concentration and recommended dilution are provided on the Control Certificate. Before use, briefly centrifuge the Positive Control to allow all of the liquid to collect in the bottom of the vial.

Calculation of Results:

1. Subtract the average background value from the test values for each sample.
   2. Using the results observed for the calibrators construct a calibration curve. The appropriate curve fit is that of a four- parameter logistics curve. A second order polynomial (quadratic) or other curve fits may also be used.
   3. Interpolate test sample values from the calibration curve. Correct for sample dilution factor to arrive at B2M concentration in original sample.
   
   1. Reliable and reproducible results will be obtained when the assay procedure is carried out with a complete understanding of the information contained in the package insert instructions and with adherence to good laboratory practice.
   2. Factors that might affect the performance of the assay include proper instrument function, cleanliness of glassware, quality of distilled or de-ionized water, and accuracy of reagent and sample pipettings, washing technique, incubation time or temperature.

Restrictions: For Research Use only

Handling

Storage: 4 °C

Storage Comment: 1. Complete Kit: The expiration date for the kit is stated on the outer label. The recommended storage temperature is 4°C. Note: See long term storage recommendations below for the Rat B2M Calibrator and Positive Control.
2. Diluent: The 5X Diluent Concentrate is stable until the expiration date. The 1X working solution is stable for at least one week from the date of preparation. Both solutions should be stored at 4°C.
3. Wash Solution: The 20X Wash Solution Concentrate is stable until the expiration date. The 1X working solution is stable for at least one week from the date of preparation. Both solutions can be stored at room temperature (RT, 16-25°C) or at 4°C.
4. Enzyme-Antibody Conjugate: Undiluted anti-B2M-HRP conjugate should be stored at 4°C and diluted immediately prior to use. The working conjugate solution is stable for up to 8 hours.
5. TMB Substrate Solution: The TMB Substrate Solution should be stored at 4°C and is stable until the expiration date.
6. Stop Solution: The Stop Solution should be stored at 4°C and is stable until the expiration date.
7. Microtiter Plate: Anti-rat B2M coated wells are stable until the expiration date and should be stored at 4°C in the sealed foil pouch with a desiccant pack.
8. Rat B2M Calibrator: Long Term Storage : Upon receipt, aliquot the calibrator and store them frozen. They will be stable until expiration date.

Expiry Date: The expiry date is stated on the label.

Target Details for beta-2 Microglobulin

Target: beta-2 Microglobulin (B2M) (beta-2-Microglobulin (B2M))

Alternative Name: Beta 2-Microglobulin (B2M Products)

Synonyms: Ly-m11 ELISA Kit, beta2-m ELISA Kit, beta2m ELISA Kit, bwm ELISA Kit, wu:fa94c05 ELISA Kit, wu:fb10a09 ELISA Kit, b2m-W01 ELISA Kit, b2m-W03 ELISA Kit, b2m-Z01 ELISA Kit, b2m-Z02 ELISA Kit, b2m-Z03 ELISA Kit, B2M ELISA Kit, b2m ELISA Kit, B2MG ELISA Kit, wu:fb66f11 ELISA Kit, wu:fj01f08 ELISA Kit, zgc:64161 ELISA Kit, b-2-m ELISA Kit, beta-2-microglobulin ELISA Kit, beta-2 microglobulin ELISA Kit, beta-2-microglobulin S homeolog ELISA Kit, beta-2-microglobulin, like ELISA Kit, B2M ELISA Kit, B2m ELISA Kit, b2m ELISA Kit, b2m.S ELISA Kit, b2ml ELISA Kit

Background: Beta 2-Microglobulin (B2M) is an 11 kDA protein. It forms the subunit of the MHC class I molecule and associates with the outer membrane of many cells including lymphocytes. It is present in low levels in serum and urine of normal people, but at a higher concentration in people with renal diseases, kidney transplants and various other inflammatory and infectious conditions.

Pathways: TCR Signaling, Regulation of Leukocyte Mediated Immunity, Positive Regulation of Immune Effector Process