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Description
Human AGGF1 ELISA KitProduct Specification Usage Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High precision pipette and gun tips: 0. 5 10uL, 5 50uL, 20 200uL, 200 1000uL 3. 37 constant temperature box 4. Distilled water or deionized water Sample processing and requirements: 1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4 overnight, then centrifuge at 1000g for 20
Product Specification
| Usage |
Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High-precision pipette and gun tips: 0.5-10uL, 5-50uL, 20-200uL, 200-1000uL 3. 37℃ constant temperature box 4. Distilled water or deionized water Sample processing and requirements: 1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4℃ overnight, then centrifuge at 1000×g for 20 minutes, and take the supernatant, or store the supernatant at -20℃ or -80℃, but avoid repeated freezing and thawing. 2. Plasma: Collect the specimen using EDTA or heparin as an anticoagulant. Centrifuge the specimen at 1000 × g for 15 minutes at 2-8°C within 30 minutes of collection. The supernatant can be assayed or stored at -20°C or -80°C, but avoid repeated freezing and thawing. 3. 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 the tissue and mince it. Add the minced tissue to the appropriate volume of PBS (generally a 1:9 weight-to-volume ratio, e.g., 1 g of tissue sample to 9 mL 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 the supernatant can be assayed. 4. 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 enzyme conjugate are sequentially added to microwells pre-coated with an Angiogenic Factor With G Patch And FHA Domains 1 (AGGF1) 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 Angiogenic Factor With G Patch And FHA Domains 1 (AGGF1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Angiogenic Factor With G Patch And FHA Domains 1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | G-patch FHA domain angiogenic factor 1 (AGGF1) is a protein encoded by the AGGF1 gene. This gene is an angiogenic factor with G-patch and forkhead-associated domains. It is primarily expressed in activated, plump endothelial cells, where it regulates angiogenesis and vascular development. AGGF1 interacts with multiple proteins involved in vascular development. Mutations in AGGF1 are associated with various cancers and can cause the rare congenital disorder Kripple-Trenauen syndrome. While AGGF1 primarily functions in endothelial cells, vascular smooth muscle cells, and osteoblasts, it is also active in mast cells, cardiac cells, Kupffer cells, and hematopoietic stem cells. AGGF1 mRNA has been detected in the heart, kidneys, and limbs, suggesting that the protein likely functions in these organs as well. | |||||||||||||||||||||||||||||||||
| 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 | Serum, plasma, tissue homogenates and other biological fluids |
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4.1 ★★★★★
Based on 18 reviews
Sort
Product Reviews
★★★★★ 1
A real review.
Color: 2PC, Color: 2PC
When I saw the Unbreakable Teddy Extreme Bear 2.0, I thought, Finally! A toy that might survive my 9-month-old Rottweiler puppy, Pepper. She’s an aggressive chewer with a love for stuffies—she adores them, right up until she disembowels them and feasts on their cotton insides like a tiny, unhinged zombie. The toy’s name, "Unbreakabear," sounded promising. Durable? Stuffie-like? For large breeds? Sign me up!
Fifteen minutes later, it looked like a crime scene from a horror movie.
The Good:
Pepper was IN LOVE with this toy... for about 10 minutes.
The double pack is nice in theory—you know, so you have a backup when the first one inevitably doesn’t live up to its "unbreakable" promise.
The Bad:
False Advertising: This toy is marketed as "extremely durable for aggressive chewers." But let me tell you: Pepper’s chewing isn’t aggressive—it’s surgical. She shredded the bear’s ears, ripped off the tail, and then went straight for the brain like she was auditioning for Zombie Dogs Gone Wild. It didn’t even put up a fight.
Durability (or Lack Thereof): Calling this toy "extreme" is like calling a paper towel roll a chew toy. One good shake, and the seams basically exploded in surrender.
Safety Concerns: Once Pepper exposed the stuffing (and the metaphorical "brains"), it was a race to see if I could stop her from swallowing it. Stuffing everywhere. The floor looked like someone had murdered a carnival bear.
The Ugly:
Walking into the room after Pepper was done was like stumbling upon the aftermath of a teddy bear massacre. The "Unbreakabear" lay limp on the floor, its ears and tail missing, stuffing spewed around like it was the victim of a bear-sized horror flick. Pepper, triumphant, sat there with bits of fluff clinging to her mouth, looking like a deranged extra from The Walking Dead.
Final Thoughts:
If your dog is a light chewer who likes to gently cuddle their toys, this might be fine. But if your dog sees toys as a challenge, run. Stick to hard rubber toys, Kongs, or something without a vulnerable, squishy middle.
As for the warranty? Sure, I could try to get a replacement, but what’s the point? Pepper will just go full zombie surgeon again.
Would I recommend this toy? Only if you’re filming a sequel to Teddy Bear Massacre. For aggressive chewers, this is nothing but fluff and disappointment.
Pepper’s Review: 5/5 stars for taste. 0/5 stars for durability.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on January 15, 2025
★★★★★ 1
Toy does not live up to its name
Color: 1PC
This product is not sturdy enough for an aggressive chewer as it is advertised. It took my 7-year-old "Rottie" approximately 15-20 minutes to have this "unbreakable, in destructive aggressive chewer" toy rip open and stuffing everywhere. A complete waste of money and I would not recommend this toy to anyone with an aggressive chewer.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on April 19, 2026
★★★★★ 1
Total Rip Off Zero Star Rating Warranted
Color: 1PC
I purchased this product for my dog because it was advertised as indestructible. The second my dog opened the package and put the bear in his mouth the “indestructible bear” was torn apart just below the bear’s head. It was useless and the stuffing was exposed. We had to take the bear from our dog and throw it away. The product was literally destroyed, rendered useless, and unusable in seconds. We had to take the product from our dog fearing he would infest the stuffing which I’m sure would have been harmful to our dog. I have never seen a dog toy that was so easily torn apart and this has never happened to any of the many toys we have purchased and/or given to our dog for play toys. I would rate this product zero stars if possible. Unfortunately one star is the lowest rating you can give when submitting your review. I’m embarrassed I spent money on what truly is junk. We and anyone else should demand a refund for purchasing this product.
Sincerely
One Disappointed and Disgusted Amazon Prime Member
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on December 27, 2024
★★★★★ 1
It’s not tough at all. Easily destroyed.
Color: 1PC, Color: 1PC
Well after 2 minutes my dog completely destroyed it.
That was a waste of money.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on June 11, 2026
★★★★★ 2
not indestructible
Color: 1PC
My dog ate through it in 4 days. The bear is not indestructible. It died a very undignified death. I am convinced that my dog was a goat in a past life. He eats everything that is not nailed down.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on April 14, 2026
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