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Description
Human ADORA2a 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 ultrasonically disrupted or repeatedly frozen and thawed. Finally, the homogenate is centrifuged at 5000×g for 5-10 minutes and the supernatant is collected for analysis. 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 an Adenosine A2a Receptor (ADORA2a) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB converts to blue under the catalysis of HRP and to yellow under the action of acid. The intensity of the color is positively correlated with the amount of Adenosine A2a Receptor (ADORA2a) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Adenosine A2a Receptor ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | The adenosine A2A receptor, also known as ADORA2A, is an adenosine receptor and also represents the gene encoding it. This protein is a member of the G protein-coupled receptor (GPCR) family. It possesses seven transmembrane alpha helices, with an extracellular N-terminus and an intracellular C-terminus. Additionally, a small alpha helix, commonly referred to as helix 8 (H8), exists on the intracellular side, close to the membrane. The crystal structure of the adenosine A2A receptor reveals a ligand-binding pocket that is distinct from other structurally determined GPCRs, namely the beta-2 adrenergic receptor and hormones. Beneath this primary binding pocket lies a secondary binding pocket. The crystal structure of A2A bound to the antagonist ZM241385 reveals that a sodium ion can be found at this location in the protein, hence its designation as the "sodium-binding pocket." This gene encodes a protein that is one of several adenosine receptor subtypes. | |||||||||||||||||||||||||||||||||
| 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 | 0.15-10 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates and other biological fluids |
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4.4 ★★★★★
Based on 17 reviews
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Product Reviews
★★★★★ 1
Nope, not for Murphy the alligator hunter! 🐊🙄
Color: grass-green, Color: grass-green
Murphy, our 75 pound lab, loves to chew. We can’t give him anything soft or even anything remotely made for casual play as he chews them to pieces- literally. So when I saw this cute little alligator, with fairly good reviews, I bought it with high expectations. Nope. It didn’t have a chance- It’s in the garbage. ☹️
First I have to comment on the eyes. I don’t know what they are made of, but the “whites” of the eyes pealed off like paint within the first few minutes Murphy had it. He wasn’t even chewing at the eyes; they just pealed off. I can’t imagine that is good for dogs to ingest, but ok, I threw the white pieces away, and gave it back to him. I do have to say that he loved chewing on it- even without adding any kind of treat to it. So we watched him chew at it for about 30-40 minutes without it being dissected into a million different parts. I was impressed.... Until I took it away from him to be played with later.... during this time he was able to tear parts of the mouth (see pictures), and while all pieces are still in tact, I cannot imagine another 30 minutes of him having this in one piece. So, while I’m very aware that Murphy has limited toys to chew on, I will continue my search for indestructible toys for his chewing pleasure.
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Reviewed in the United States on April 9, 2021
★★★★★ 4
Cute toy maybe not for aggressive chewers
Color: grass-green
I bought this because I have an 3 1/2 yr old golden doodle. She tears every toy apart within half hour of having it. The only toys she hasn’t is Kong toys.
This is a cute toy. She was shredding pieces off the rubber with half hour of chewing it:(
I thought it was very similar to the material of Kong toys?
She loves chewing it and playing with it but I probably would not buy it for really aggressive large dogs who tear up toys.
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Reviewed in the United States on March 31, 2026
★★★★★ 5
Durable.
Color: grass-green
Very durable. My dog loves it.
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Reviewed in the United States on April 30, 2026
★★★★★ 3
My lab loved it too much, too fast…
Color: blue, Color: blue
It arrived an hour ago and my lab has it in pieces within minutes. It kept him busy for a bit though! I’ll give it to him again but I’ll have to sit with him, for sure. No squeak? I guess he just didn’t get to that part of the toy. It felt sturdy but ultimately didn’t pass the test.
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Reviewed in the United States on January 5, 2026
★★★★★ 5
Dog Toy for aggressive dog
Color: yellow, Color: yellow
I give this dog toy a big WOW! The material is made out of great hard rubber material.
My Doggie tends to rip through most of the dog toys. The size was perfect. The material the hardness easy to use. I’m gonna buy more for Christmas as presents. The weight was perfect. She loves it.I love it.
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Reviewed in the United States on May 22, 2026