PAPPA ELISA Kit

Catalog No. ABIN5680718

Product Details PAPPA ELISA Kit

Target: PAPPA (Pregnancy-Associated Plasma Protein A, Pappalysin 1 (PAPPA))

Reactivity: Mouse

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Detection Range: 0.24 ng/mL - 10.3 ng/mL

Minimum Detection Limit: 0.24 ng/mL

Application: ELISA

Purpose: The Mouse PAPP-A enzyme linked immunosorbent assay (ELISA) kit provides materials for the quantitative measurement of PAPP-A in mouse serum, cell supernatant, and other biological fluids.

Sample Type: Plasma, Serum

Analytical Method: Quantitative

Sensitivity: 0.02 ng/mL

Components:

• Mouse PAPP-A Calibrator A/Sample Diluent
• Mouse PAPP-A Calibrator B-F
• Mouse PAPP-A Controls
• PAPP-A Antibody Coated Microtitration Strips
• PAPP-A Assay Buffer:
• PAPP-A Conjugate Diluent
• Mouse PAPP-A Biotin Conjugate Concentrate
• Mouse PAPP-A Streptavidin-Enzyme Conjugate Ready-to-Use
• TMB Chromogen Solution
• Wash Concentrate A
• Stopping Solution

Material not included:

1. Microtitration plate reader capable of absorbance measurement at 450nm, 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.

Application Details

Sample Volume: 25 μL

Assay Time: 3.5 h

Plate: Pre-coated

Reagent Preparation:

1. Mouse PAPP-A Calibrators B-F and Mouse PAPP-A Controls I & II: Tap and reconstitute Mouse PAPP-A Calibrator B-F and Mouse PAPP-A Controls I & II each with 1 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. Mouse PAPP-A Biotin Conjugate Solution: The mouse PAPP-A Biotin Conjugate Concentrate should be diluted at a ratio of 1 part into 50 parts of the PAPP-A conjugate diluent, according to the number of wells used. For an entire plate, pipet exactly 220 μL of the Biotin Conjugate Concentrate into 11 mL of the PAPP-A Conjugate Diluent. NOTE: The Biotin conjugate concentrate should be freshly diluted 10-30 minutes prior to use.
4. 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.

Sample Collection:

• 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. 15

Assay Procedure:

Allow all samples and reagents to reach room temperature. Mix reagents thoroughly by gentle inversion before use. Calibrators, controls and samples should be assayed in duplicate. NOTE: All serum samples reading higher than the highest calibrator should be thoroughly mixed and diluted in the 0 ng/mL (Calibrator A) prior to assay.

1. Mark the microtitration strips to be used.
2. Pipet 25 μL of the calibrators, controls and samples to the appropriate wells.
3. Add 50 μL of the PAPP-A Assay Buffer to each well using a precision pipette.
4. Incubate the wells, shaking at 600-800 rpm on an orbital microplate shaker, for 2hrs at room temperature (23±2 °C).
5. During the last 20-30 minutes of incubation, prepare the Biotin Conjugate Solution by diluting the Mouse PAPP-A Biotin Conjugate Concentrate in PAPP-A Conjugate Diluent as described under the Preparation of the Reagents section of this package insert.
6. Aspirate and wash each well 5 times with the wash solution using an automatic microplate washer.
7. Add 100 μL of the biotin conjugate solution to each well using a precision pipette.
8. Incubate the wells, shaking at 600-800 rpm on an orbital microplate shaker, for 1hr at room temperature (23±2 °C).
9. Aspirate and wash each well 5 times with the wash solution using an automatic microplate washer.
10. Add 100 μL of the Streptavidin-Enzyme Conjugate-RTU to each well using a repeater pipette.
11. Incubate the wells, shaking at 600-800 rpm on an orbital microplate shaker, for 30 minutes at room temperature (23±2 °C).
12. Aspirate and wash each well 5 times with the wash solution using an automatic microplate washer.
13. Add 100 μL of the TMB chromogen solution to each well using a precision pipette. Avoid direct exposure to heat and sunlight.
14. Incubate the wells, shaking at 600-800 rpm on an orbital microplate shaker, for 8-10 min at room temperature (23±2 °C). NOTE: Visually monitor the color development to optimize the incubation time.
15. Add 100 μL of the stopping solution to each well using a precision pipette.
16. 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.

Calculation of Results:

NOTE: The results in this package insert were calculated by plotting the data on a log vs. log scale using a cubic regression curve-fit. Other data reduction methods may give slightly different results.

1. Calculate the mean OD for each calibrator, Control, or Unknown.
2. Plot the log of the mean OD readings for each of the Calibrators along the y-axis versus log of the PAPP-A concentrations in ng/mL along the x-axis, using a cubic regression curve-fit.
3. Determine the PAPP-A concentrations of the unknowns from the calibration curve by matching their mean OD readings with the corresponding PAPP-A concentrations.
4. Any sample reading higher than the highest calibrator should be appropriately diluted using Mouse PAPP-A Calibrator A and re-assayed.
5. Any sample reading lower than the analytical sensitivity should be reported as such.
6. If a high reading sample is diluted, multiply the value by the appropriate dilution factor.

Restrictions: For Research Use only

Handling

Precaution of Use: The following precautions should be observed: a) Follow good laboratory practice. b) Use personal protective equipment. Wear lab coats and disposable gloves when handling immunoassay materials. c) Handle and dispose of all reagents and material in compliance with applicable regulations. WARNING: Potential Biohazardous Material This reagent may contain some animal and/or human source material (e.g. serum) or materials used in conjunction with human source materials. Handle all reagents and patient samples at a Biosafety Level 2, as recommended for any potentially infectious human material in the Centers for Disease Control/National Institutes of Health manual "Biosafety in Microbiological and Biomedical Laboratories," 5th Edition, 20071. WARNING: Potential Chemical Hazard Some reagents in this kit contain Pro-Clean 400 and Sodium azide2 as a preservative. Pro-Clean 400 and Sodium Azide in concentrated amounts are irritants to skin and mucous membranes. For further information regarding hazardous substances in the kit, please refer to the MSDS.

Storage: 4 °C

Target Details for PAPPA

Target: PAPPA (Pregnancy-Associated Plasma Protein A, Pappalysin 1 (PAPPA))

Alternative Name: PAPP-A (PAPPA Products)

Synonyms: 8430414N03Rik ELISA Kit, IGFBP-4ase ELISA Kit, PAG1 ELISA Kit, PAPP-A ELISA Kit, ASBABP2 ELISA Kit, DIPLA1 ELISA Kit, PAPA ELISA Kit, PAPPA1 ELISA Kit, Pappa ELISA Kit, pappalysin 1 ELISA Kit, pregnancy-associated plasma protein A ELISA Kit, PAPPA ELISA Kit, Pappa ELISA Kit, Pappa1 ELISA Kit