IGFBP5 ELISA 试剂盒

• IGFBP-4 Calibrators A
• -- Cal-127F IGFBP-5 Calibrators B - F (Lyophilized)
• IGFBP-5 Controls I & II
• IGFBP-5 Coated Microtitration strips
• IGFBP-5 Assay Buffer
• IGFBP-5 Antibody-Enzyme Conjugate RTU
• TMB Chromogen Solution
• Stopping Solution
• Wash Concentrate A

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

1. IGFBP-5 calibrators B-F and IGFBP-5 Controls I & II: Tap and reconstitute IGFBP-5 Calibrator B-F and IGFBP-5 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 (23 ± 2 °C) 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 tube 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 specimens and reagents to reach room temperature (23 ± 2 °C) and mix thoroughly by gentle inversion before use. Calibrators, controls, and unknowns should be assayed in duplicate. Note: Do not dilute the calibrators and controls. All samples reading higher than the highest calibrator should be diluted in the 0 ng/mL Calibrator A prior to assay.

1. Reconstitute IGFBP-5 Calibrator B-F and IGFBP-5 Controls I & II each with 0.5 mL deionized water as mentioned in the Preparation of Reagents section. Solubilize for 10 minutes, Mix well by gentle vortex.
2. Label the microtitration strips to be used.
3. Pipette 20 μL of the Calibrator, Controls and Unknowns to the appropriate wells
4. Add 100 μL of the IGFBP-5 Assay Buffer to each well using a repeater pipette.
5. Incubate the plate, shaking at a fast speed (600-800 rpm) on an orbital microplate shaker, for 60 minutes at room temperature (23 ± 2 °C).
6. Aspirate and wash each strip 5 times with Wash Solution (350 μL/per well) using an automatic microplate washer.
7. Add 100 μL of the IGFBP-5 Enzyme Conjugate Solution to each well using a repeater pipette.
8. Incubate the plate, shaking at a fast speed (600-800 rpm) on an orbital microplate shaker, for 60 minutes at room temperature (23 ± 2 °C).
9. Aspirate and wash each strip 5 times with the Wash Solution (350 μL/per 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.
13. 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 IGFBP-5 concentration on X- axis using a cubic regression curve-fit. Alternatively, log vs. log quadratic regression curve-fit can be used. 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 optical density (OD) for each calibrator, Control, or Unknown.
3. Plot the log of the mean OD readings for each of the Calibrators along the y-axis versus log of the IGFBP-5 concentrations in ng/mL along the x-axis, using a cubic regression curve-fit.
4. Determine the IGFBP-5 concentrations of the Controls and unknowns from the calibration curve by matching their mean OD readings with the corresponding IGFBP-5 concentrations.
5. Any sample reading higher than the highest Calibrator should be appropriately diluted with the 0 ng/mL (CAL A / Sample Diluent) and re- assayed.
6. Any sample reading lower than the analytical sensitivity should be reported as such.