ELISA Mouse - NGAL/LCN2

ELISA is the most commonly used method of detecting and quantifying the concentration of an antigen in an unknown sample. During the experiment, If you get a weak signal, then make sure reagents are at room temperature before starting the assay. Try increasing incubation times to ensure maximal antibody binding and amplify the signal. Secondly, if you get values above 0 in the negative control indicates a high background signal. Try to consider reducing your antibody concentration and prevent non-specific binding of antibodies by using affinity-purified antibody and suitable blocking buffers. To avoid high well to well variation, do not stack plates during incubation, no bubbles in the plate and wash wells thoroughly to avoid variation.

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Found 3 matching solutions for this experiment

Upstream tips
-Bring all reagents to room temperature before use. Allow all components to sit for a minimum of 15 minutes with gentle agitation after initial reconstitution.
-Working dilutions should be prepared and used immediately.
Protocol tips
-A standard curve should be generated for each set of samples assayed.
-It is suggested to start Reagent Diluent optimization for serum and plasma samples by using PBS supplemented with 10-50% animal serum.
-Do not use buffers with animal serum to reconstitute or dilute the Detection Antibody or Streptavidin-HRP B.
Upstream tips
-Store at 4°C for 6 months, at -20°C for 12 months.
-Avoid multiple freeze-thaw cycles.
-The standard solutions are best used within 2 hours.
Protocol tips
-The diluted Avidin-Biotin-Peroxidase Complex (ABC) solution should not be prepared more than 1 hour prior to the experiment.
-Prepare the substrate solution immediately before use or bring the pre-made substrate to room temperature.
-Sodium azide is an inhibitor of HRP. Do not include the azide in buffers or wash solutions if HRP-labeled conjugate is used for detection.
Upstream tips
-Equilibrate all reagents to room temperature (18-25°C) prior to use.
Protocol tips
-Avoid foaming or bubbles when mixing or reconstituting components.
-Ensure plates are properly sealed or covered during incubation steps.
-Complete removal of all solutions and buffers during wash steps is necessary to minimize background.
Downstream tips
-Unused well strips should be returned to the plate packet and
stored at 4°C.
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