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| Catalog # | Availability | Size | Quantity | Unit Price | Save For Later Wish List | |
|---|---|---|---|---|---|---|
| EKFT10107HU-96 | 2 weeks | 96T | $525.00 |
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| EKFT10107HU-48 | 2 weeks | 48T | $370.00 |
Product Overview | ||||||||||||||||||||||||||||
| Name | Human hepatitis A virus-Immunoglobulin G (HAV-IgG) ELISA Kit | |||||||||||||||||||||||||||
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| Reactivity | Human | |||||||||||||||||||||||||||
| Sensitivity | Qualitative | |||||||||||||||||||||||||||
| Assay Range | Qualitative | |||||||||||||||||||||||||||
| Detection Method | Indirect ELISA | |||||||||||||||||||||||||||
| Sample type(s) | Serum, Plasma, Cell Culture Supernatant, cell or tissue lysate, Other liquid samples | |||||||||||||||||||||||||||
| Principle of the Assay | ||||||||||||||||||||||||||||
| This kit was based on indirect ELISA. HAV-Ag was pre-coated onto 96-well plates. The test samples were added to the wells, unbound conjugates were washed away with wash buffer. Then added HRP- Conjugates, if there were any HAV -IgG in the samples, it would form a HAV-Ag- HAV-IgG- HRP- Conjugates complex. TMB substrates were used to visualize HRP enzymatic reaction. It was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The optical density of developed color is read with a suitable photometer at 450nm with a selected reference wavelength within 650 nm. | ||||||||||||||||||||||||||||
| Sample Collection and Storage | ||||||||||||||||||||||||||||
| Isolate the test samples soon after collecting, then, analyze immediately (within 2 hours). Or aliquot and store at -20°C for long term. Avoid multiple freeze-thaw cycles. Serum: Coagulate the serum at room temperature (about 1 hours). Centrifuge at approximately 1000 × g for 15 min. Analyze the serum immediately or aliquot and store at -20°C. Plasma: Collect plasma with heparin or EDTA as the anticoagulant. Centrifuge for 15min at 2-8°C at 1500 x g within 30 min of collection. For eliminating the platelet effect, suggesting that further centrifugation for 10 min at 2-8°C at 10000 x g. Analyze immediately or aliquot and store frozen at -20°C. Note: Samples to be used within 3 days may be stored at 2-8°C, otherwise samples must be stored at -20°C (≤1 month) or -80°C (≤2 months) to avoid loss of bioactivity and contamination. Hemolyzed samples are not suitable for use in this assay. Wash Buffer Preparation: Dilute 20mL of Concentrated Wash Buffer to 800 mL of Wash Buffer with deionized or distilled water. | ||||||||||||||||||||||||||||
| Calculation of Results | ||||||||||||||||||||||||||||
| Negative control: A ≤0.1 (If one well A>0.1, it should be abandoned; If two wells A>0.1, it should be retested) Positive control: A ≥ 0.8 Calculation of the Cutoff Value Cutoff Value = NCx × 2.1 NCx: Mean Absorbance of Negative Control. When NCx<0.05, Calculate as 0.05. Note: Sample with absorbance values ≤ Cutoff Value is NON-REACTIVE and are considered NEGATIVE for HAV-IgG. Sample with absorbance values > Cutoff Value are considered POSITIVE for HAV-IgG. Positive samples are recommended to be diluted 1000 times with normal saline to repeat the experiment and determine the results again. | ||||||||||||||||||||||||||||
| Kit Components | ||||||||||||||||||||||||||||
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| Precautions for Use | ||||||||||||||||||||||||||||
| 1.After opening and before using, keep plate dry. 2. Before using the Kit, balance the reagents at room temperature at least 30 mins. 3. Storage TMB reagents avoid light. 4. Washing process is very important, not fully wash easily cause a false positive. 5. Don’t let Micro plate dry at the assay, for dry plate will inactivate active components on plate. 6. Don’t reuse tips and tubes to avoid cross contamination. 7. Avoid using the reagents from different batches together. | ||||||||||||||||||||||||||||
| Material Required But Not Supplied | ||||||||||||||||||||||||||||
| 1.Microplate reader (wavelength: 450nm) 2.37°C incubator 3.Automated plate washer 4.Precision single and multi-channel pipette and disposable tips 5.Clean tubes and Eppendorf tubes 6.Deionized or distilled water | ||||||||||||||||||||||||||||
| Manual Washing | ||||||||||||||||||||||||||||
| Discard the solution in the plate without touching the side walls. Clap the plate on absorbent filter papers or other absorbent material. Fill each well completely with 350ul wash buffer and soak for 1 to 2 minutes, then aspirate contents from the plate, and clap the plate on absorbent filter papers or other absorbent material. Repeat this procedure two more times for a total of THREE washes. | ||||||||||||||||||||||||||||
| Automated Washing | ||||||||||||||||||||||||||||
| Aspirate all wells, then wash plate THREE times with 350ul wash buffer. After the final wash, invert plate, and clap the plate on absorbent filter papers or other absorbent material. It is recommended that the washer be set for a soaking time of 1 minute. | ||||||||||||||||||||||||||||
| Assay Procedure | ||||||||||||||||||||||||||||
| 1.Wash plate 2 times before adding sample and control wells. 2. Label the sample wells, 2 Negative Controls, 1 Positive Control and 1 blank well. 3. Add 50μL sample, Negative Controls and Positive Control to each well (except blank well) and gently tap the plate to ensure thorough mixing. Seal the plate with a cover and incubate at 37°C for 30 min. 4. Remove the cover, and wash plate 5 times with Wash buffer and let the wash buffer stay in the wells for 1 minute each time. 5. Add 50 µL HRP- Conjugates into each well (except blank well) and gently tap the plate to ensure thorough mixing. 6. Seal the plate with a cover and incubate at 37°C for 30 min. 7. Remove the cover, and wash plate 5 times with Wash buffer and let the wash buffer stay in the wells for 1 minute each time. 8. Add 50 μl of TMB substrate A and 50 μl of TMB substrate B into each well (except blank well). Gently tap the plate to ensure thorough mixing. Cover the plate and incubate at 37°C in dark for 10 min. And the shades of blue can be seen in the Positive Controls. Negative Controls wells show no obvious color. 9. Add 50 μl of Stop solution into each well and mix thoroughly. The color changes into yellow immediately. 10. Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution. (Use the blank well to set zero) | ||||||||||||||||||||||||||||
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