PMN Elastase ELISA Assay

$935.00

The PMN Elastase ELISA Assay Kit is intended for the quantification of human PMN Elastase in EDTA or citrate plasma, exudate, bronchoalveolar lavage fluid, cerebrospinal fluid and seminal plasma. The Eagle Biosciences PMN Elastase ELISA Assay Kit is for research use only and should not be used for diagnostic procedures.

SKU: ARG80507 Categories: ,

PMN Elastase ELISA Assay

PMN Elastase ELISA Assay is For Research Use Only

Size: 1×96 wells
Sensitivity: 0.2 ng/ml
Dynamic Range: 15.6-1000 ng/ml
Incubation Time: 2.5 hours
Sample Type: Plasma, Seminal Plasma Exudate, Bronchoalveolar Lavage Fluid, and Cerebrospinal Fluid
Sample Size: 100 µL
Alternative Names: Neutrophils


Assay Principle

The PMN Elastase ELISA Assay employs the quantitative sandwich enzyme immunoassay technique. An antibody specific for PMN Elastase has been pre-coated onto a microtiter plate. Standards or samples are pipetted into the wells and any PMN Elastase present is bound by the immobilized antibody. After washing away any unbound substances, a HRP-conjugated antibody specific for PMN Elastase is 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 PMN Elastase 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 PMN Elastase in the sample is then determined by comparing the O.D of samples to the standard curve.


Related Products

PMN-Elastase (Serum/Plasma) ELISA Assay
PMN-Elastase (Stool) ELISA Assay Kit
Pancreatic Elastase ELISA Kit

Additional Information

Assay Background


The human organism reacts with an inflammatory response to attacks of invading pathogens (micro-organisms and viruses) or damaged tissue (after accidents or surgery). Polymorphonuclear (PMN) granulocytes play an important role as primary defense cells in this inflammatory reaction. Different bloodstream mediators (cytokines, leukotrienes, complement factors, bacterial endotoxins, clotting and fibrinolysis factors) attract and stimulate these cells to phagocytize and destroy not naturally occurring agents.
PMN granulocytes use proteinases to digest these agents and tissue debris. One of these proteinases is PMN elastase which is localized in the azurophilic granules of the polymorphonuclear granulocytes. During phagocytosis of foreign substances these enzymes are also partially excreted into the extracellular surrounding, where the activity of PMN elastase is regulated by inhibitors (esp. the α1-proteinase inhibitor, α1-PI). An overwhelming release of PMN elastase, however, can exceed the inhibitory potential of the α1-proteinase inhibitor. Thus, enzymatically active PMN elastase, together with simultaneously produced oxidants (O2-radicals, H2O2, OH-radicals), can cause local tissue injury. Due to the bloodstream and lymphatic system, however, α1-PI is delivered subsequently and eventually able to form a complex with all excreted elastase. Therefore, the concentration of the PMN elastase/ α1-PI complex correlates with the released PMN elastase and can be used as a measure for the activity of granulocytes during an inflammatory response. Primarily, determinations of PMN elastase find its application in observation of the course of trauma, shock and sepsis. Further indications are the areas of hemodialysis, infections by obstetrics, joint diseases, and effusions of sport injuries, intestinal affection, pancreatitis, cystic fibrosis and male adnex affections.

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, controls and samples in duplicate into wells.
    3. Cover the wells and incubate for 1 hour at RT.
    4. Aspirate each well and wash, repeating the process 3 times for a total 4 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 150 μl of Enzyme-conjugated Antibody to each well. Cover wells and incubate for 1 hour at RT.
    6. Aspirate each well and wash as step 4.
    7. Add 200 μl of TMB Reagent to each well. Incubate for 20 minutes at room temperature in dark.
    8. Add 50 μl of Stop Solution to each well.
    9. Read the OD with a microplate reader at 450 nm immediately.

 

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