Fumonisin ELISA Kit

Catalog No. ABIN997086

Product Details

Target: Fumonisin

Reactivity: Chemical

Detection Method: Colorimetric

Method Type: Competition ELISA

Application: ELISA

Purpose: Fumonisin quantitative test is based on the principle of the enzymelinked immu-nosorbent assay.

Analytical Method: Quantitative

Sensitivity: 0.5 ng/mL

Material not included: 1.50, 100 and 1000 μL micropipettes
2. ELISA reader (450 nm)
3. Grinder
4. Centrifuge Reagents 1. Methanol (80 %)

Application Details

Plate: Pre-coated

Assay Procedure:

1. Prepare samples as described above.
   2. Pipette 100 μL standards or prepared samples in duplicate into the appropriate wells of the microtiter plate. Immediately add 50 μL fumonisin-peroxidase conjugate and 50 μL anti-fumonisin antibody into each well.
   3. Cover the microtiter plate with a plastic foil and incubate for 60 minutes at room temperature.
   4. Wash the plate three times as follows: Discard the contents of the wells (dump or aspirate). Pipette 300 μL of diluted washing solution into each well. After the third repetition empty the wells again and remove residual liquid by striking the plate against a paper towel. The wash procedure is critical. Insufficient washing will result in poor precision and falsely elevated absorbencies.
   5. Pipette 100 μL of substrate solution into each well.
   6. Allow the reaction to develop in the dark (e.g. cupboard or drawer, the chromogen is light-sensitive) for
   2. minutes at room temperature.
   7. Stop enzyme reaction by adding 100 μL of stop solution (0.5 M H2SO4) into each well. The blue colour will turn yellow upon addition.
   8. After thorough mixing, measure absorbance at 450 nm (reference wavelength 620 nm), using an ELISA reader. The color is stable for 30 minutes. Grind approximately 50-100 g of maize or grain sample to a fine powder. Extract 3 g ground sample with 9 mL 80 % methanol in distilled water on a shaker for at least 15 minutes. Clear the sample by centrifugation (10 min, 2000 g) or filtration. Dilute cleared sample 1:15 in sample diluent (e.g. 100 μL cleared sample + 1.4 mL sample diluent).

Calculation of Results:

1. Calculate the average optical density (OD 450 nm) for each set of reference standards or samples.
   
   2. Construct a standard curve by plotting the mean optical density obtained for each reference standard against its concentration in ng/mL on semi-log graph paper with the optical density on the vertical (y) axis and the concentration on the horizontal (x) axis.
   
   3. Using the mean optical density value for each sample, determine the corresponding concentration of fumonisin in ng/mL from the standard curve. Depending on experience and/or the availability of computer capability, other methods of data reduction may be employed.
   4. The diluted samples must be further converted by the appropriate dilution factor (45 for the above described extraction). The factor is dependent on the sample preparation procedure employed. Typical Standard Values The following table contains an example for a typical standard curve. The binding is calculated as percent of the absorption of the 0 ng/mL standard. These values are only an example and should not be used instead of the standard curve which has to be measured in every new test. Fumonisin (ng/mL) ( % binding of 0 ng/mL) 0 100 0.5 87 2 62 5 38 10 23 25 11
   

Restrictions: For Research Use only

Handling

Storage: 4 °C

Storage Comment: Store at 2-8 °C

Target Details

Target: Fumonisin

Target Type: Chemical