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Data sheet
| Validity | 12 months from manufacture date |
| Assay Range | Qualitative: Positive & Negative Control |
| Assay principle | The Filaria IgG4 ELISA consists of one enzymatically amplified sandwich-type immunoassay. Positive, negative and low positive control samples are provided to maintain the kit’s integrity. In this assay, control samples and unknown serum samples are diluted into the Filaria Sample Dilution Buffer, then incubated in microtiter wells which have been coated with IWb123 (1-6), followed by incubation with anti-human IgG4 antibody labeled with the enzyme horseradish peroxidase (HRP). After the incubation and a washing step, the wells are incubated with a tetramethylbenzidine (TMB) substrate. An acidic stopping solution is then added and the degree of enzymatic turnover of the substrate is determined by absorbance measurement at 450 nanometers. The absorbance measured is directly proportional to the concentration of specific IgG4 antibodies to IWb123 present. A set of positive and negative controls are provided as internal controls. These are provided to monitor the integrity of the kit components. |
| Sample type | 10µl Serum |
| INTENDED USE | The Filaria IgG4 ELISA is for qualitative detection of specific IgG4 antibodies in specimens to highly specific antigen IWb123 target antigen expressed primarily in infective stage larvae (L3) of the lymphatic-dwelling parasite Wuchereria bancrofti (Wb). This product is for research use only. Not for use in diagnostic procedures. |
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MATERIALS AND COMPONENTS The Filaria IgG4 ELISA kit contains sufficient reagents for one plate of 96 wells (8 x12 strips) each. The kit contains the following reagents: Warning: Do not use any reagents where damage to the packaging has occurred.
1. FILARIA ANTIGEN COATED MICROTITER STRIPS: Strip holder in Ziploc foil, containing 96 polystyrene microtiter wells coated with IWb123 in each well. Stable at 2-8°C until the expiration date.
2. FILARIA SAMPLE DILUTION BUFFER: Two bottles, 25 ml, Tris-HCl buffered solution (pH 7.2-7.6) with Tween 20 (0.05%), preservative (0.05% proclin-300) and additives. Use for the dilution of test samples, positive and negative controls. This sample dilution buffer is also used for the elution of dried blood spots as described in the Dry Blood Reference guide below. Stable at 2-8°C until the expiration date.
3. FILARIA NEGATIVE CONTROL: One vial, 50 μl. Negative serum. The Negative Control will aid in monitoring the integrity of the kit. Store at 2-8oC.
4. FILARIA LOW POSITIVE CONTROL: One vial, 50 μl. Low reactive positive sample. The Low Positive Control will aid in monitoring the integrity of the kit. Store at 2-8ºC.
5. FILARIA POSITIVE CONTROL: One vial, 50 μl. Positive serum containing <1.0% BSA and <0.05% Proclin 300. The Positive Control will aid in monitoring the integrity of the kit. Store at 2-8ºC.
6. 100X ENZYME CONJUGATE FOR FILARIA: One vial, 150ul, contains Mouse monoclonal anti-human IgG4 conjugated with horseradish peroxidase (HRP) in Tris buffered solution (pH~7.4), preservative and additives. Store at 2-8⁰C until the expiration date.
7. CONJUGATE DILUENT FOR FILARIA: One bottle, 14ml contains PBS buffered solution (pH~7.4) with preservative and additives. The 100X Enzyme Conjugate is diluted into this Conjugate Diluent before use. Stable at 2-8°C until expiration date.
8. 10X WASH BUFFER: One bottle, 120 ml, of 10X concentrate of phosphate buffered saline with Tween 20 (pH 6.8-7.0). Stable at 2-8°C until the expiration date.
9. LIQUID TMB SUBSTRATE: One bottle, 12ml, ready to use. Contains 3, 3’, 5, 5’-tetramethylbenzidine (TMB) and hydrogen peroxide in a citric-acid citrate buffer (pH 3.33.8). Stable at 2-8°C until the expiration date.
Note: The TMB substrate should always be stored in the lightprotected bottle provided.
10. STOP SOLUTION: One bottle, 6 ml, of ready to use 1N Sulfuric Acid. Used to stop the enzymatic turnover of the substrate. Stable at 2-8°C until the expiration date.
Ensuring Assay Validity: The results on the table below must be obtained using provided positive and negative controls to calculate discrimination capacity of the assay. Non-fulfillment of these criteria is an indication of deterioration of reagents or an error in the test procedure and the assay must be repeated.
Kit controls (positive, low positive and negative control) are provided to assess ONLY the kit integrity.
The Discrimination Capacity is defined as the ratio of the mean Positive Control OD450 to the mean Negative Control OD450 (PC ÷ NC).
Interpretation of the results:
1. Samples with spectrophotometric readings greater than the geographically established Cut-off value are considered to be “Reactive” and samples below this criterion are considered to be “Non-Reactive”.
2. Any “Reactive” sample must be repeated to verify the result. Values near the Cut-off are considered to be doubtful and the assay must be repeated in triplicate to establish the sample status.