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Description
Human ADAM17 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. Cell Lysis Buffer: Gently wash adherent cells with ice-cold PBS, then trypsinize and collect cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be collected directly by centrifugation. Wash collected cells three times with ice-cold 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 freeze-thaw cycles or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and remove the supernatant for analysis. 5. Cell Culture Supernatant: Centrifuge at 1000×g for 20 minutes. Remove the supernatant for analysis or store at -20°C or -80°C, avoiding repeated freeze-thaw cycles. 6. 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 capture antibody against A disintegrin and metalloprotease 17 (ADAM17). 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 A disintegrin and metalloprotease 17 (ADAM17) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human A Disintegrin And Metalloprotease 17 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | disintegrin-metalloproteinase 17 (ADAM17), also known as TACE (tumor necrosis factor-α converting enzyme), is a 70 kDa enzyme that belongs to the ADAM protein family of disintegratins and metalloproteinases. It is involved in processing tumor necrosis factor-α (TNF-α) on the cell surface and removing it from the intracellular membrane of the trans-Golgi network. This process, also known as "shedding," involves the cleavage and release of a soluble ectodomain from the membrane-bound proprotein (such as pro-TNF-α) and is of known physiological importance. | |||||||||||||||||||||||||||||||||
| 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 homogenate, cell lysate, cell culture supernatant and other biological fluids |
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4.8 ★★★★★
Based on 576 reviews
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Product Reviews
★★★★★ 5
No breakup, very sweet, instalove
Format: Kindle
Omegaverse and doesn't disappoint! Sweet guys, newly Omega FMC. The boyfriends are boyfriends. What's not to love? No angst, no breakup.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on February 23, 2025
★★★★★ 4
Pretty Darn Good
Format: Kindle
So I’ve been on a omega kick and this definitely hit the spot. Madison was frustrating at times with how she acted towards Lucas, Gray, and Rian. It was like she said towards the end, she didn’t believe she deserved nice things.
It would have been nice to hear from her best friends again. They kind of were there in the beginning and the gone except for mention of text messages received from them. I feel like her friends would have been great help in encouraging Madison to go with the pack and never give Brent another chance because he was toxic.
I loved Rian. His personality was awesome. His humor. His ability to make Madison comfortable whenever she was feeling overwhelmed. And the fact he fell for her and she fell for him first. They are cute together.
I do feel like Lucas was the odd man out though. Like Lucas didn’t develop as much of a relationship with Madison. I would have really liked to see more development in the relationship between them. It was also the same with him and Rian. There is really no relationship displayed. Most of the relationship being displayed is between Rian and Gray.
Nevertheless, I loved reading about the dynamic that came to fruition during the entirety of this story. Madison finally got her happiness. And Brent finally got punched in the face. Everyone got exactly what they deserve.
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Reviewed in the United States on September 6, 2022
★★★★★ 3
Pack's Promise was okay but not great
Format: Kindle
Pack's Promise was okay but not great. I won't recommend it to anyone that I know.
PRO:
* Very likable characters
* Lots of steamy scenes that are written very well
* The spelling and grammar are good
* The punctuation is good with the exception of using hyphens instead of commas. Lots of hyphens. Lots and lots of hyphens.
CON:
* Almost no interactions with any characters outside of Madison and the pack
* Nearly no plot. They meet, get together for a heat, agree to make it permanent, done
* Quite a few typos such as extraneous words, missing words and words out of order
THINGS TO KNOW:
* More steamy scenes than storytelling
* A lot of MM & MMM, some MFMM during heat
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Reviewed in the United States on January 5, 2023
★★★★★ 5
such a good read
Format: Kindle
Madison, Lucas, Grey and Rian were made for each other!!! First time reading from this author and I’m not disappointed!!! Absolutely love the Love in this book and couldn’t ask for a better OV!
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Reviewed in the United States on October 25, 2023
★★★★★ 4
Fluffy and Nice Omegaverse
Format: Kindle
… this would have made 5 stars but for 2 reasons.
A.) there were quite a few typos; misspelled words, missing quotations, “the his” mistakes, and various signs that maybe a proofread would do good.
B.) the writing was quite textbook. Late blooming omega is struggling with her new self, finds a absurdly wealthy pack of alphas, every thing is almost insta-love but she resists, then decides to love herself and let everyone be happy.
Rian was my favourite (obviously the author’s favourite too because he got the most page time) but I wish we could see more of his CEO side? He went to work maybe ONCE the entire time. Gray was supposed to be the “growly one” but he turned out to be puppy dog. Lucas was a genius brainiac doctor - but also super alpha with an aggressive hindbrain with a breeding k*nk?? And then there was no actual “breeding”??
Spice 3/5 - normally omegaverse books are super high on messy smut but this was tamer.
Romance 3/5 - insta-love that was then resisted because of personal hangup’s
Plot 2/5 - weird paced head hopping, showing the same scene from different POV’s that made me feel like it was 2 steps backward, 1 step forward.
Humour 4/5 - there were a dozen lines that genuinely made me chuckle out loud
Would have been five stars but the lack of proofreading and the predictable plot made me unable to get up to ADORED IT level - four stars is still and official ENJOYED IT, y’all. This isn’t a bad rating. The “Club Heat” has intriguing possibilities so I’m going to give the second one a shot.
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Reviewed in the United States on March 31, 2023