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Description
Human WNT5A ELISA KitProduct Specification Usage Required experimental equipment: 1. Microplate reader (450nm) 2. High precision pipettes and pipette tips: 0. 5 10uL, 5 50uL, 20 200uL, 200 1000uL 3. 37C incubator 4. Distilled or deionized water Sample preparation and requirements: Tissue homogenization: Rinse the tissue with pre chilled PBS (0. 01M, pH 7. 4) to remove residual blood (lysed red blood cells in the homogenate will affect the measurement results). Weigh and
Product Specification
| Usage | Required experimental equipment: 1. Microplate reader (450nm) 2. High-precision pipettes and pipette tips: 0.5-10uL, 5-50uL, 20-200uL, 200-1000uL 3. 37°C incubator 4. Distilled or deionized water Sample preparation and requirements: Tissue homogenization: Rinse the tissue with pre-chilled PBS (0.01M, pH 7.4) to remove residual blood (lysed red blood cells in the homogenate will affect the measurement results). Weigh and mince the tissue. Add the minced tissue to the appropriate volume of PBS (generally a 1:9 weight-to-volume ratio, e.g., 1g of tissue sample to 9mL of PBS. The specific volume can be adjusted according to experimental needs and recorded. It is recommended to add protease inhibitors to the PBS) in a glass homogenizer and grind thoroughly on ice. To further lyse tissue cells, the homogenate can be sonicated or repeatedly frozen and thawed. Finally, centrifuge the homogenate at 5000×g for 5-10 minutes, and collect the supernatant for analysis. Cell Lysis Buffer: Gently wash adherent cells with pre-chilled PBS, then trypsinize and collect the cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be collected directly by centrifugation. Wash the collected cells three times with pre-chilled PBS and resuspend in 150-200 μL of PBS per 1×10^6 cells (it is recommended to add protease inhibitors to the PBS; if the cell count is very low, reduce the PBS volume appropriately). Disrupt the cells by repeated freezing and thawing or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and collect the supernatant for analysis. Cell Culture Supernatant: Centrifuge at 1000×g for 20 minutes. Collect the supernatant for analysis, or store at -20°C or -80°C, but avoid repeated freezing and thawing. Other biological fluids: Centrifuge at 1000xg for 20 minutes, remove the supernatant, and test. Pre-test preparation: 1. Remove the test kit from the refrigerator 10 minutes in advance and equilibrate to room temperature. 2. Prepare the standard gradient working solution: Add 1 mL of universal diluent to the lyophilized standard, let it stand for 15 minutes to completely dissolve, then gently mix (concentration is 10 ng/mL). Then dilute to the following concentrations: 10 ng/mL, 5 ng/mL, 2.5 ng/mL, 1.25 ng/mL, 0.625 ng/mL, 0.3125 ng/mL, 0.15625 ng/mL, and 0 ng/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each. Pipette 500uL of the 10ng/mL standard working solution into the first EP tube and mix thoroughly to make a 5ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube serves as a blank well; there is no need to pipette liquid from the penultimate tube. See the figure below for details. 3. Preparation of biotinylated detection antibody working solution: Centrifuge the concentrated biotinylated antibody at 1000×g for 1 minute 15 minutes before use. Dilute the 100× concentrated biotinylated antibody to a 1× working concentration with universal diluent (e.g., 10uL concentrate + 990uL universal diluent). Prepare and use immediately. 4. Prepare the enzyme conjugate working solution: 15 minutes before use, centrifuge the 100× concentrated enzyme conjugate at 1000×g for 1 minute. Dilute the 100× concentrated HRP enzyme conjugate to a 1× working concentration with universal diluent (e.g., 10 μL of concentrate + 990 μL of universal diluent). Prepare immediately. 5. Prepare the 1× wash solution: Dispense 10 mL of 20× wash solution into 190 mL of distilled water (concentrated wash solution removed from the refrigerator may crystallize; this is normal. Allow to stand at room temperature until the crystals have completely dissolved before preparing). Procedure: 1. Remove the desired strips from the aluminum foil bag after equilibration at room temperature for 10 minutes. Seal the remaining strips in a ziplock bag and return to 4°C. 2. Sample addition: Add 100 μL of sample or standard of varying concentrations to the corresponding wells. Add 100 μL of universal diluent to the blank wells. Cover with a film and incubate at 37°C for 60 minutes. (Recommendation: Dilute the sample to be tested at least 1-fold with universal diluent before adding it to the ELISA plate. This will reduce the impact of matrix effects on the test results. The sample concentration should be multiplied by the corresponding dilution factor when calculating the final sample concentration. It is recommended to run replicates for all test samples and standards.) 3. Add Biotinylated Antibody: Remove the ELISA plate and discard the liquid without washing. Add 100 μL of Biotinylated Antibody Working Solution directly to each well. Cover with a film and incubate at 37°C for 60 minutes. 4. Wash: Discard the liquid and add 300 μL of 1x Wash Solution to each well. Let stand for 1 minute, shake off the wash solution, and pat dry on absorbent paper. Repeat this process three times (a plate washer can also be used). 5. Add Enzyme Conjugate Working Solution: Add 100 μL of Enzyme Conjugate Working Solution to each well. Cover with a film and incubate at 37°C for 30 minutes. 6. Washing: Discard the liquid and wash the plate five times as in step 4. 7. Adding substrate: Add 90 μL of substrate (TMB) to each well, cover with a sealing film, and incubate at 37°C in the dark for 15 minutes. 8. Adding stop solution: Remove the ELISA plate and add 50 μL of stop solution directly to each well. Immediately measure the OD value of each well at a wavelength of 450 nm. Calculating experimental results: 1. Calculate the average OD value of the standard and sample replicates and subtract the OD value of the blank well as a correction factor. Plot the standard curve of the four-parameter logistic function on double-logarithmic graph paper, with concentration as the horizontal axis and OD value as the vertical axis. 2. If the sample OD value is higher than the upper limit of the standard curve, dilute the sample appropriately and retest. Multiply the sample concentration by the corresponding dilution factor. |
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| Theory | This kit uses a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Sample, standard, biotin-labeled detection antibody, and HRP conjugate are sequentially added to microwells pre-coated with a Wingless Type MMTV Integration Site Family, Member 5A (WNT5A) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by HRP peroxidase and to yellow by acid. The intensity of the color is positively correlated with the amount of Wingless Type MMTV Integration Site Family, Member 5A (WNT5A) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Wingless Type MMTV Integration Site Family, Member 5A ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Wingless-type MMTV integration site family member 5A, also known as Wnt-5a protein, is encoded by the WNT5A gene. Disorders associated with WNT5A include Robinaud syndrome, autosomal dominant 1, and autosomal dominant Robinaud syndrome. Pathways associated with this gene include the canonical Wnt pathway and CDK-mediated phosphorylation and clearance of Cdc6. Gene ontology annotations associated with this gene include DNA-bound transcription factor activity and protein domain-specific binding. An important homolog of this gene is WNT5B. | |||||||||||||||||||||||||||||||||
| General Notes | 1. Strictly adhere to the specified incubation time and temperature to ensure accurate results. All reagents must be at room temperature (20-25°C) before use. Refrigerate reagents immediately after use. 2. Improper plate washing may result in inaccurate results. Ensure that all liquid in the wells is aspirated thoroughly before adding substrate. Do not allow the wells to dry out during incubation. 3. Remove any residual liquid and fingerprints from the bottom of the plate, as this will affect the OD value. 4. The substrate developer solution should be colorless or very light in color. Do not use substrate solution that has turned blue. 5. Avoid cross-contamination of reagents and specimens to prevent erroneous results. 6. Avoid direct exposure to strong light during storage and incubation. 7. Do not expose any reagents to bleaching solvents or the strong fumes emitted by bleaching solvents. Any bleaching agent will destroy the biological activity of the reagents in the kit. 8. Do not use expired products, and do not mix components with different product numbers and batches. 9. Recombinant proteins from sources other than the kit may not be compatible with the antibodies in this kit and will not be recognized. 10. If there is a possibility of disease transmission, all samples should be managed properly and samples and testing devices should be handled according to prescribed procedures. |
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| Storage Temp. | If the unopened kit is stored at 4°C, the shelf life is 6 months. | |||||||||||||||||||||||||||||||||
| Test Range | 0.156-10 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, cell culture supernatants, and other biological fluids |
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4.2 ★★★★★
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Product Reviews
★★★★★ 5
Love these pillows!
Size: Queen (2 pack)
I’m a side and stomach sleeper. I’ve bought so many pillows recently trying to find one that doesn’t kill my neck or shoulder. I felt like I was looking for a unicorn. For some reason this pillow just works. It has a higher loft, which I prefer, but it’s also super squishy, which I also prefer. It somehow has the best of both worlds. They feel high quality, are adjustable in terms of density since you can remove as much filling as you want, they actually fill my pillowcases so they’re not flopping around the pillow, and they maintain their shape after a full night of sleeping on them, the price point for two pillows is just right, and best of all, they actually stay cool. I am a hot sleeper so that last point is worth the cost of entry all by itself. I have no neck or shoulder pain after getting a full night’s sleep on these pillows. The only negative is the initial smell of the pillows. It’s pretty strong, but I’m sure this will lessen over time.
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Reviewed in the United States on April 3, 2026
★★★★★ 5
Soft and cool.
Color: Cooling Blue Cover, Size: Queen (Pack of 2)
Love this pillow set. Comfortable addition and nice and cool at all times. Going to be great for when the hot weather hits
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Reviewed in the United States on June 8, 2026
★★★★★ 5
As advertised for once! Lol
Color: Cooling Blue Cover, Size: Queen (Pack of 2), Color: Cooling Blue Cover, Size: Queen (Pack of 2)
Queen size set of 2 pillows, I'm giving it 5stars first bc it has a good length and width!! If you've ever bought memory foam pillows before you know some are small!! Anyways after having it for 5nights, we loved that me and my huddy could adjust the Thickness amount we had in it to our needs, me needing a thick pillow and my husband needing a flatter one. So this was perfect for that. So far its nice to sleep on, stays cool, doesnt get hot. I would say its on the frimer side but softer the more foam you take out! Really its just good for the amount you spend!! Will it last 10years probably not but what does now a days. This is the only time my huddy n I bought the same pillow and both love it so I'm just still in shock it worked out so well!!
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Reviewed in the United States on April 28, 2026
★★★★★ 5
Adjustable pillows
Color: Cooling Blue Cover, Size: Queen (Pack of 2)
I love these because they don't go flat. You can pull out whatever you need to make them comfy for you. They come in two different layers. So you have that protective cover over the main case that holds the memory foam. They're amazing to sleep on. I have had these last for years without going flat. If you hate flat pillows this is the way to go. They are very easy to wash. You just have to separate layers. They are very thick, you can adjust the thickness. Everyone in my house has at least one. They are worth the money. I have never had to replace them, because they went flat.
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Reviewed in the United States on May 6, 2026
★★★★★ 4
Good pillows
Color: Cooling Blue Cover, Size: Queen (Pack of 2)
Takes some getting used to but and the cooling doesn’t last very long in my opinion, but overall amazing comfy pillows. The materials are good and are washable. I mostly stopped waking up with a stiff neck and back.
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Reviewed in the United States on April 20, 2026