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Description
Human CYP24A1 ELISA KitProduct Specification Usage Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High precision pipette and pipette 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: Tissue, homogenates, cell lysates and other biological fluids Preparation before testing: 1. Please take out the test kit from the refrigerator 10 minutes in
Product Specification
| Usage |
Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High-precision pipette and pipette 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: Tissue, homogenates, cell lysates and other biological fluids Preparation before testing: 1. Please take out the test kit from the refrigerator 10 minutes in advance and equilibrate to room temperature. 2. Prepare a gradient standard working solution: Add 1 mL of universal diluent to the lyophilized standard. Let stand for 15 minutes to completely dissolve, then gently mix (concentration 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 500 μL of universal diluent to each tube. Pipette 500 μL of the 10 ng/mL standard working solution into the first EP tube and mix thoroughly to make a 5 ng/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 Cytochrome P450 24A1 (CYP24A1) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by peroxidase (HRP) catalysis and to yellow by acid. The intensity of the color is positively correlated with the amount of Cytochrome P450 24A1 (CYP24A1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Cytochrome P450 24A1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Cytochrome P450 family 24, subfamily A, member 1 (abbreviated as CYP24A1), also known as vitamin D₃ 24-hydroxylase, is a member of the cytochrome P450 superfamily and is encoded by the CYP24A1 gene. It is a mitochondrial monooxygenase that catalyzes the 24-hydroxylation of calcitriol (1,25-dihydroxyvitamin D₃). Conditions associated with CYP24A1 include hypercalcemia, infantile dysregulation, and Brody disease. Pathways associated with this gene include metabolism and metabolism of fat-soluble vitamins. Gene ontology annotations associated with this gene include oxidoreductase activity and heme binding. An important homologue of this gene is CYP27A1. | |||||||||||||||||||||||||||||||||
| 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 | Tissuehomogenates, celllysates and otherbiologicalfluids |
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4.5 ★★★★★
Based on 18 reviews
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Product Reviews
★★★★★ 5
Puppy
Pattern Name: Heartbeat Sheep Grey
Simply the best! These hold up and are a puppy favorite!
They did change the heartbeat sound and it is a bit much for the newborn puppies.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on December 13, 2025
★★★★★ 1
Pillow is Unsafe
My eleven month old Cavalier, Lacey, loved this heartbeat pillow for about two hours. She carried it everywhere. Then she located the velcro opening. She ripped it open, pulled out the stuffing. I stuffed the heart in the bottom but I cannot locate it in the pillow nor have I found it anywhere in my home. Velcro closure is not enough to keep a curious puppy from figuring out how to open the pillow.
Would I recommend this product: No I do not recommend this item. It was a total waste of my hard earned $. The closure needs to be reworked/improved. As it is now, do not leave your puppy alone with this heartbeat pillow because it is not safe!
The only positive is it kept Lacey from barking for two hours while she played with it. I cannot attest that it would have helped her severe separation anxiety and yelping every time I leave the house.
Lynne Weston
Amazon Customer
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Reviewed in the United States on July 14, 2024
★★★★★ 5
Good value
Size: Large
So far so good! Seems sturdy enough for my little guy. Good value for the price.
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Reviewed in the United States on June 2, 2026
★★★★★ 5
my dogs love these! their new constant companions!
When I first opened the box and saw these three octopuses, I thought they looked a little anemic, but looks can definitely be deceiving because my dogs love them.
We try to be really careful about the toys we buy because one of my dogs swallowed a squeaker in the past and had to have surgery. Ever since then, we've been really mindful about what toys they have and making sure we're watching them when they play.
I have to say these have been great. My dogs carry them everywhere. They take them outside, bring them back inside, and I've had to wash them multiple times because they are so loved and so well used. For some reason, my dog who is a constant chewer and destroyer of toys has not chewed through these. They feel relatively tough, especially considering how lightweight they are. They do have a squeaker inside, but they also have something crinkly in them that makes noise when the dogs are running around and playing with them. My dogs seem to love that. My dogs also look super cute running around with the octopus legs hanging out of their mouths. Fun product!
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Reviewed in the United States on June 9, 2026
★★★★★ 5
Great toys for dogs
I received these octopus toys for my dog because he destroys stuffed toys almost immediately, and I liked that these were no stuffing. They ended up being a big hit right away. The squeakers kept him interested longer than most toys do, and he loves shaking the octopus legs around while playing. Since there’s no stuffing inside, I don’t have to worry about coming home to fluff all over the house, which has honestly been a huge plus. The toys are lightweight and easy for him to carry around, and they work well for toss and fetch too. I have a medium sized dog, but I could see these being fun for smaller or even larger dogs depending on how aggressive of a chewer they are. They seem decently durable for plush toys. Mine have held up better than expected considering how rough my dog can be with his toys. The stitching has stayed together pretty well so far. I also like that the pack comes with three because one is always getting hidden somewhere in the house. It’s nice having extras ready to go. These have definitely helped keep my dog entertained and busy, especially on days when we can’t spend as much time outside.
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Reviewed in the United States on May 28, 2026