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Description
Mouse ACAT1 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: Adherent cells should be gently washed with pre-chilled PBS, then trypsinized and harvested by centrifugation at 1000×g for 5 minutes. Suspension cells can be harvested directly by centrifugation. Collected cells should be washed three times with pre-chilled PBS and resuspended 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. 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 acetyl-CoA acetyltransferase, mitochondrial (ACAT1) 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 acetyl-CoA acetyltransferase, mitochondrial (ACAT1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Mouse | |||||||||||||||||||||||||||||||||
| Synonym | Mouse Acetyl-CoA acetyltransferase, mitochondrial ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Acetyl-CoA acetyltransferase 1 (ACAT1), also known as acetoacetyl-CoA thiolase (ACAT), is an enzyme encoded by the ACAT1 gene. It is an acetyl-CoA C-acetyltransferase. This gene encodes a mitochondrially localized enzyme that catalyzes the reversible formation of acetyl-CoA from two molecules of acetyl-CoA. The ACAT1 enzyme has several unique properties. First, it is activated by potassium ion binding near the CoA binding site and the catalytic site. This binding results in structural changes in the active site loop. Furthermore, the enzyme can use 2-methyl-branched acetoacetyl-CoA as a substrate, making it a unique thiolase. It is regulated at both the transcriptional and translational levels. Enhanced ACAT1 enzyme activity is transcriptionally promoted by leptin, angiotensin II, and insulin in human monocytes/macrophages. Mutations in this gene are associated with a deficiency of the encoded mitochondrial acetoacetyl-CoA thiolase protein, also known as ketothiolase deficiency. Its expression has been associated with the manifestation of prostate cancer. | |||||||||||||||||||||||||||||||||
| 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, and other biological fluids |
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4.8 ★★★★★
Based on 627 reviews
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Product Reviews
★★★★★ 5
Cane Corso approved....
Size: Large, Pattern Name: Pack of 1
My Cane Corso loves this ball!!! She carries it around in her mouth like a round pacifier.
Sometimes I'll wake up with the ball next to my head on my pillow. Lol
This is the third one I've bought.
She doesn't destroy them at all . Somehow she loses them. Yesterday she had two floating around. Lol.
Earlier I gave her one of those hard rubber balls with feet... She destroyed it within minutes. She can skin a tennis ball and chew it in half in about 5 minutes.
This ball is by far a favorite toy of her's. And she doesn't chew it to bits .
And it glows in the dark which is fun for playing fetch at night.
It's very easy to clean.
Sometimes I put little snacks inside for her while she's laying down with her ball.
I will definitely keep buying these balls everytime we lose one . They make her happy. And that makes me happy.
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Reviewed in the United States on May 12, 2026
★★★★★ 5
BEST GLOW BALL! MY COLLIE BLUE HEELER MIX IS OBSESSED
Size: Large, Pattern Name: Pack of 1
Well if I could just say...Mommas a fan too....BUT I think my dog should be the sponsor of CHUCKIT because he owns everything and he BREATHES, DREAMS, RUNS with #chuckit balls!! Since 8wks. old & Now 10 yrs young, BDAY=8.10.26....he wakes up with it in his mouth and falls asleep with IT! I HAVE THEEEE PROOF LOL SERIOUSLY OBSESSED! #chuckit #makemydogyoursponsor WE HAVE EVERY BALL X10 FOR SOME, launchers, bucket o balls , you name it so yeah #obssesedisanunderstatement #bordercollieblueheelermix #HARLEY ♡ nice colors and the durability is amazing 👏 know if you have a chewer I'm not sure because he just likes to catch the ball and then chew it sometimes and then leave it alone, until next time around. But the GLOW BALL just being cewed for so long and the textire is obviously different so it does end up cracking and you throw it away, definitely different rubber but highly recommend them both actually ALL of them👌🎉🥳🫶🏼 my Sisters 2 yr old pit destroyed them in minutes but only FOR PLAY , so IF YOU DONT WANT THAT BALL chewed to pcs. Put it up LOL and oh
all of the balls EXCEPT the GLOW BALL stay true to form and durability, for real Harleys had some of them for years!! And nothing wrong with the CHUCKIT ball 10☆☆☆☆☆☆☆☆☆☆ SIZE is perfect too he love the medium 2.5" ball. Perfect for catching for hrs on end. You also have a choice you can get squeaky, crunchy,or nothing or glow in the dark, 3 sizes to fit your pupper..
.. so many choices and it doesn't have to be a ball you can pick from a ton of toys ' tuggers ,launchers and indoor toys too. As for the Chew resistance...mm touchy...like I said. Aggressive chewer are more likely to tear it up way quicker than the just playing mainly like I want to fetch dog. Either way. I dont purchase any other brand besides chuck it. Has always my pick, like 20+ yrs now lol. Hands down. This is not a paid thing or promoted or whatever. Just felt i should review for chuck it♡ to date has made my dog happy and healthier for years to come♡ thanks
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Reviewed in the United States on May 8, 2026
★★★★★ 4
These are great!
Size: Medium, Pattern Name: Pack of 2
My dog loves these and now won't play fetch with any other ball. I love that they glow bright so we can play fetch after I get home from work (it gets dark around 5 now). The dog and I both like that they whistle. They also squish easily and he loves to chomp on them while he runs around. Only reason I took off a star is because he chewed a piece of it off and now it doesn't whistle or fly as far. That being said, it seems to stand up well to all the chomping he does on it. I should not have left him alone with it, I think he held it down and pulled on it. If your dog usually makes it his mission to destroy things I wouldn't leave him alone with it, other than that I absolutely recommend these.
Update 12/12/20: I got another dog last year and he also loves these balls. Probably the favorite ball for both dogs, they go into hyperdrive when they see them. New dog tends to be more destructive than the older one and I still don't leave them unattended with these balls, but new dog has not destroyed one. I made the mistake of leaving one in the Chuckit overnight once. Ball was wet from playing in the snow and became lopsided. Now doesn't fly very well but my weird dog still likes it. It is also starting to dry out and crack. None of the other glow balls have done this, I'm assuming it is also from leaving it in the Chuckit overnight.
Other than that, these balls are great. I did read a few reviews that said charging them with UV light made them stay brighter for a longer period, but I did not notice a difference in my quick test of this theory. Just keep a good flashlight with you and brighten them up as needed.
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Reviewed in the United States on December 6, 2018
★★★★★ 5
Good for night sessions
Size: Medium, Pattern Name: Pack of 1
Picked this up for evening fetch sessions and it’s been a win. It charges up fast, couple minutes under a lamp or in sunlight and it’s good to go, and the glow is genuinely bright, not the sad dim purple some of these put out. Makes it way easier to find in the yard at dusk, and the dog can actually track it.
It’s a little on the softer side compared to a regular tennis ball, but durable enough to hold up to my big dog, who is not gentle with toys. Still kicking after plenty of chomping and slobber. Fits a standard Chuckit! launcher perfectly, which is the whole point.
Pros:
• Glows brightly and clearly in the dark
• Charges fast from any light source
• Holds up well even with a big, enthusiastic chewer
• Fits standard ball-throwing sticks
• Great for evening or early-morning fetch
Cons:
• A bit softer than a regular tennis ball — a determined power-chewer could probably destroy it eventually
• Glow fades after a while, so you’ll need to recharge it during long sessions
• Only glows if it’s been charged, which is obvious but easy to forget
Tips:
• Charge it under a bright light or in the sun for a few minutes before heading out
• A flashlight works in a pinch if you forgot to charge it
• Keep it out of permanent fetch duty — rotate with a regular ball so it lasts longer
• Not a chew toy, even though it’s tempting to leave it with the dog
Five stars. If you’ve got a dog and a yard, this makes evening fetch actually doable.
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Reviewed in the United States on May 24, 2026
★★★★★ 5
Westies Fav
Size: Small, Pattern Name: Pack of 1
Best Westie ball ever, my dog loves this ball. Exactly her size! 3" tennis balls are too big & she rips the fur off. this silicone-ish ball has no fur & is 2&1/2 "
We have 2" balls from her electronic ball laucher, but she still loves this glow in the dark best!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on May 4, 2026
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