beta Defensin-4 ELISA Assay Kit

$895.00

The Eagle Biosciences beta Defensin-4 ELISA assay kit is intended for the quantification of Human BD-4 / beta Defensin-4 in serum, plasma, cell culture supernatants The Eagle Biosciences beta Defensin-4 ELISA assay kit is for research use only and should not be used for diagnostic procedures.

SKU: ARG80902 Categories: , ,

beta Defensin-4 ELISA Assay Kit

For Research Use Only

Size: 1×96 wells
Sensitivity: 0.8 pg/ml
Dynamic Range: 1.56-100 pg/ml
Incubation Time: 4 hours
Sample Type: Serum, Plasma, Cell culture Supernatants
Sample Size: 100 µL

Additional Information

Assay Background

Defensins form a family of microbicidal and cytotoxic peptides made by neutrophils. Members of the defensin family are highly similar in protein sequence. This gene encodes defensin, beta 4, an antibiotic peptide, which is locally regulated by inflammation.

Assay Principle

The Eagle Biosciences beta-Defensin-4 ELISA Assay Kit is for the quantification of BD-4/β-Defensin-4 in serum, plasma and cell culture supernatant. This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for BD-4/β-Defensin-4 has been pre-coated onto a microtiter plate. Standards or samples are pipetted into the wells and any BD-4/β-Defensin-4 present is bound by the immobilized antibody. After washing away any unbound substances, a biotin conjugated antibody specific for BD-4/β-Defensin-4 and the Streptavidin-HRP Enzyme complex are added to each well and incubate. After washing away any unbound antibody-enzyme reagent, a substrate solution (TMB) is added to the wells and color develops in proportion to the amount of BD-4/β-Defensin-4 bound in the initial step. The color development is stopped by the addition of acid and the intensity of the color is measured at a wavelength of 450 nm ±2 nm. The concentration of BD4/β-Defensin-4 in the sample is then determined by comparing the O.D of samples to the standard curve.

Assay Procedure

1. Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal it.

2. Add 100 μl of standards, samples and zero controls (standard diluent buffer) into wells. Incubate for 1.5 h at 36 °C.

3. Prepare 1X Antibody conjugate 20 minutes before use.

4. Aspirate each well and wash, repeating the process four times for a total five washes. Wash by filling each well with 1× Wash Buffer (350 μl) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating, decanting or blotting against clean paper towels.

5. Add 100 μl 1X Antibody conjugate into each well. Cover wells and incubate for 1 hour at 36°C.

6. Prepare 1X HRP-Streptavidin solution 20 minutes before use. (Protect from light)

7. Aspirate each well and wash as step 4.

8. Add 100 μl of 1X HRP-Streptavidin solution to each well. Cover wells and incubate for 30 minutes at 36°C.

9. Aspirate each well and wash as step 4.

10. Add 100 μl of TMB Reagent to each well. Incubate for 15 minutes at 36°C in dark.

11. Add 100 μl of Stop Solution to each well. The color of the solution should change from blue to yellow.

12. Read the OD with a microplate reader at 450nm immediately.

Manual

Product Manual