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Description
Human NLRP3 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 20 ng/mL). Then dilute to the following concentrations: 20 ng/mL, 10 ng/mL, 5 ng/mL, 2.5 ng/mL, 1.25 ng/mL, 0.625 ng/mL, 0.3125 ng/mL, and 0 ng/mL. Serial dilution method: Take 7 EP tubes and add 500 μL of universal diluent to each tube. Pipette 500 μL of the 20 ng/mL standard working solution into the first EP tube and mix thoroughly to make a 10 ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube serves directly as a blank well; there is no need to aspirate the liquid from the penultimate tube. See the figure below for details. 3. Preparation of Biotinylated Antibody Working Solution: 15 minutes before use, centrifuge the concentrated biotinylated antibody at 1000×g for 1 minute. Dilute the 100× concentrated biotinylated antibody to a 1× working concentration using universal diluent (e.g., 10µL concentrate + 990µL universal diluent). Prepare immediately before use. 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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| Sensitivity | 0.18 ng/mL | |||||||||||||||||||||||||||||||||
| Theory | This kit utilizes 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 NLR Family, Pyrin Domain Containing Protein 3 (NLRP3) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB converts to blue under the catalysis of HRP peroxidase and to yellow under the action of acid. The intensity of the color is positively correlated with the amount of NLR Family, Pyrin Domain Containing Protein 3 (NLRP3) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human NLR Family, Pyrin Domain Containing Protein 3 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | NLR family pyrin domain-containing 3 (NLRP3) (formerly known as NACHT, LRR, and PYD domain-containing protein 3 [NALP3] and cryopyrin) is a protein encoded by the NLRP3 gene located on the long arm of chromosome 1. NLRP3 is primarily expressed in macrophages and, as a component of the inflammasome,436 detects products of damaged cells, such as extracellular ATP and crystallized uric acid. Activated NLRP3, in turn, triggers an immune response. Mutations in the NLRP3 gene have been associated with several organ-specific autoimmune diseases. | |||||||||||||||||||||||||||||||||
| 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 | 0.312-20 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, cell culture supernatants, and other biological fluids |
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4.4 ★★★★★
Based on 756 reviews
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Product Reviews
★★★★★ 5
Best toy, the end
Style: Sport, Size: 12in, Style: Sport, Size: 12in
Combined with the Max Glow ball, this is the best dog toy available on modern planet Earth.
I have a burly 92lb. pit bull mix with highly developed prey drive, and she will chase the glow ball for hours if I let her.
The tiny sport launcher (Sport 12M) is the best, because you can still throw the ball really far, but you can also slam the ball down a few feet away from yourself so it bounces up in the air and your dog has to jump to catch it, or you can throw it up really high so it bounces 15' away or so and your dog can jump to catch it. We sometimes have to play fetch in a pretty small area, so it's nice to have the small launcher for these games. The larger launchers aren't as fun in small areas. You can play the same games but it's trickier to throw the ball correctly.
Also, the small launcher is a lot easier to carry. The small launcher fits in my dog-walk-stuff backpack (with the ball in the launcher so I don't have to dig for it later), but the big ones don't easily. The launcher also has a hole in the end so you can hang it from your bag or whatever with a carabiner. The big ones are too long for that in most cases, and swing back and forth.
Also-also my dog loves to ask for the launcher, then when I give it to her she plays keep-away and it's adorable. The big launcher I have is harder for her to balance in her mouth, so she just lays down and chews on it until it gets taken away. She's chewed on all of the four Chuck Its we've owned, but she's only broken one (a big one), but that's because she got ahold of it when no one was home. The little one in my bag is totally chewed up but still works great. I think they're sturdier than the big ones. Except for the launcher she chewed into pieces, which was our fault, the only reason we've had to replace anything was because we lost it. Usually we keep the launcher and ball in the backpack so she can't get to them and so they don't get lost, so we rarely have to replace them. There is just not a better value in toys, and the startup price is really low.
Once again I really recommend the glow ball! It doesn't get soggy no matter how wet the grass or the dog mouth is. It is fun to chase because it is slippery and it collapses when it's squeezed, so it pops out of the dog's mouth and flies away if they bite it at the wrong angle; it's really bouncy, and stays bouncy because it can't be popped; it whistles so you can usually hear it even if you can't see it; and it really glows enough for nighttime catch, even if you only have your cellphone flashlight to charge it up!
UPDATE:
I should have said, we have the 12M launcher, which means the handle is 12 inches long and it uses a medium-sized ball. The medium ball is 2.5 inches in diameter, so the launcher will fit regular tennis balls, too! But my dog won't play with regular tennis balls anymore because they aren't anywhere near as fun as the glow balls, and I think she doesn't like having gross sloshy muddy tennis balls in her mouth.
I lost my short launcher awhile back and could only get a long one locally to replace it, which reminded me that you have to lean over a lot further to use the short launcher. So if you have trouble leaning over to the point that your hand is a foot above the ground, this may be hard for you to use.
I wish Amazon would let us choose colors, because my old one was green, and the big one is orange, both of which are bright enough to see in the grass from far away, even at night.
Still, best toy!
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Reviewed in the United States on May 19, 2017
★★★★★ 5
Highly recommend.
Style: Sport, Size: 12in
Love this. Dog sitting for a dog that loves to chase balls but my shoulder doesn’t like throwing them. Got this yesterday and it’s perfect. Real arm/shoulder saver.
Seems sturdy and I was concerned that a standard tennis ball would not work. We tend to lose balls when walking because this lab insists on carrying the ball in its mouth. They work.
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Reviewed in the United States on March 22, 2026
★★★★★ 5
These are awesome and I couldn't play ball with my Dobermans without it!
Style: Sport, Size: 12in
I LOVE Chuckit ball launchers. We've been using for a around 17 years and we've only had to replace one. We have 2 active, energetic Dobermans and I throw like a girl and there is now way I can throw a ball very far. With the Chuckit, I can throw more than 80 feet! My dogs need a lot of exercise and using this ball thrower allows me to make that happen. Bonus is I dont have to pick up a snobbery ball :)
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Reviewed in the United States on April 13, 2026
★★★★★ 4
Not a long shot
Style: Sport, Size: 12in
This item is much smaller than I thought it would be .
The ball is full sized . But the thrower is very small.
Works okay. But does not throw the ball very far . It’s okay for me . My dogs are small. My yard is too .
But , this is not going to throw balls really far . Like the long ones do
Thsnks
Eddie
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Reviewed in the United States on March 26, 2026
★★★★★ 5
Perfect addition
Style: Sport, Size: 12in
This is item that I needed but didnt know it. Its been great for saving my sholder anad my dog loved the longer chase to get the ball. Easy to use and lots of fun
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Reviewed in the United States on May 18, 2026