alpha Fetoprotein ELISA 试剂盒

• AFP Calibrator A/Sample Diluent
• AFP Calibrators B-F (Lyophilized)
• AFP Controls (Lyophilized)
• Anti-AFP Antibody Coated Microtitration Strips
• AFP Assay Buffer
• AFP Antibody Enzyme Conjugate Ready-To-Use
• TMB Chromogen Solution
• Wash Concentrate A
• Stopping Solution

1. Microtitration plate reader capable of absorbance measurement at 450 nm, 405nm and 630 nm.
2. Microtitration orbital plate shaker.
3. Microtitration plate washer.
4. Semi-automated/manual precision pipette to deliver 2-250 μL.
5. Semi-automated/manual precision pipette to deliver 1-10 μL.
6. Repeater Pipette.
7. Vortex mixer.
8. Deionized water.

1. AFP Calibrators B-F and AFP Controls I & II: Tap and reconstitute AFP Calibrator B-F and AFP Controls I & II each with 0.5 mL deionized water. Solubilize, mix well and use after reconstitution.
2. Wash Solution: Dilute wash concentrate 25-fold with deionized water. The wash solution is stable for one month at room temperature when stored in a tightly sealed bottle.
3. Microtitration Wells: Select the number of coated wells required for the assay. The remaining unused wells should be placed in the resealable pouch with a desiccant. The pouch must be resealed to protect from moisture.

• Serum is the recommended sample type.
• Sample handling, processing, and storage requirements depend on the brand of blood collection tube29 that you use. Please reference the manufacturer's instructions for guidance. Each laboratory should determine the acceptability of its own blood collection tubes and serum separation products.
• Samples may be stored at 4 °C if assayed within 24 hours, otherwise samples must be stored at -20 °C or -80 °C to avoid loss of bioactivity and contamination.
• Avoid assaying lipemic, hemolyzed or icteric samples.
• Avoid repeated freezing and thawing of samples. Thaw samples no more than 3 times.
• For shipping, place specimens in leak proof containers in biohazard specimen bags with appropriate specimen identification and test requisition information in the outside pocket of the biohazard specimen bag. Follow DOT and IATA requirements when shipping specimens.

Allow all samples and reagents to reach room temperature. Mix reagents thoroughly by gentle inversion before use. After reconstitution of reagents, mix thoroughly, avoiding foam. Calibrators, controls and samples should be assayed in duplicate.

1. Reconstitute AFP Calibrator B-F and AFP Controls I & II each with 0.5 mL deionized water. Solubilize for 10 minutes, Mix well.
2. Mark the microtitration strips to be used.
3. Pipet 25 μL of the calibrators, controls and unknown samples to the appropriate wells.
4. Add 50 μL of the AFP Assay Buffer to each well using a repeater pipette.
5. Incubate the wells, shaking at 600-800 rpm on an orbital microplate shaker, for 30 mins at room temperature (23 ± 2 °C).
6. Aspirate and wash each well 5 times with the wash solution (350 μL/well) using an automatic microplate washer.
7. Add 100 μL of the Antibody-enzyme conjugate ready to use solution to each well using a precision pipette.
8. Incubate the wells, shaking at 600-800 rpm on an orbital microplate shaker, for 30 mins at room temperature (23 ± 2 °C).
9. Aspirate and wash each well 5 times with the wash solution (350 μL/well) using an automatic microplate washer.
10. Add 100 μL of the TMB chromogen solution to each well using a repeater pipette. Avoid exposure to direct sunlight.
11. Incubate the wells, shaking at 600-800 rpm on an orbital microplate shaker, for 8-12 min at room temperature (23 ± 2 °C). NOTE: Visually monitor the color development to optimize the incubation time.
12. Add 100 μL of the Stopping solution to each well using a repeater pipette. Read the absorbance of the solution in the wells within 20 minutes, using a microplate reader set to 450 nm. NOTE: Zero calibrator should be programmed as "Blank" while reading the optical density. If instrument has a wavelength correction, set the instrument to dual wavelength measurement at 450 nm with background wavelength correction at 630 nm.

NOTE: The results in this package insert were calculated by plotting the log optical density (OD) data on the y-axis and log AFP concentration on X-axis using a cubic regression curve-fit. Other data reduction methods may give slightly different results.

1. Optimum results can be obtained at incubation temperature of (23 ± 2 °C).
2. Calculate the mean OD for each calibrator, control or diluted sample.
3. Plot the log of the mean OD readings for each of the Calibrators along the y-axis versus log of the AFP concentrations in ng/mL on the x-axis, using a cubic regression curve fit.
4. Determine the AFP concentrations of the controls and samples from the calibration curve by matching their mean OD readings with the corresponding AFP concentrations.
5. Any sample reading higher than the highest calibrator should be appropriately diluted using calibrator A/sample diluent and re-assayed.
6. Any sample reading lower than the analytical sensitivity should be reported as such.