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Description
Human DDC 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 handling and requirements: The detection range of the kit is not equivalent to the concentration range of the analyte in the sample. Before the experiment, it is recommended to estimate the analyte concentration in the
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 handling and requirements: The detection range of the kit is not equivalent to the concentration range of the analyte in the sample. Before the experiment, it is recommended to estimate the analyte concentration in the sample based on relevant literature and conduct preliminary experiments to determine the actual concentration in the sample. If the analyte concentration in the sample is too high or too low, dilute or concentrate the sample appropriately. If the sample type is not listed in the instructions, it is recommended to conduct a preliminary experiment to verify the validity of the test. Serum: Collect whole blood in a serum separator tube and place it at room temperature for 2 hours or at 2-8°C overnight. Then centrifuge at 1000×g for 20 minutes and remove the supernatant. Alternatively, store the supernatant at -20°C or -80°C, but avoid repeated freezing and thawing. Plasma: Collect the sample using EDTA or heparin as an anticoagulant. Within 30 minutes of collection, centrifuge at 1000×g for 15 minutes at 2-8°C. Remove the supernatant and test it. Alternatively, store the supernatant at -20°C or -80°C, but avoid repeated freezing and thawing. Tissue homogenate: Rinse the tissue with pre-chilled PBS (0.01M, pH 7.4) to remove any residual blood (lysed red blood cells in the homogenate may affect the test 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., 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 or in a homogenizer. To further lyse tissue cells, the homogenate can be sonicated or repeatedly freeze-thawed. Finally, centrifuge the homogenate at 5000×g for 5-10 minutes and remove the supernatant for analysis. Cell culture supernatant: Centrifuge at 1000×g for 20 minutes. Remove the supernatant and analyze it immediately, or store it at -20°C or -80°C, but avoid repeated freeze-thaw cycles. Cell lysis buffer: Gently wash adherent cells with ice-cold PBS, then trypsinize and harvest the cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be harvested 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 (protease inhibitors are recommended; if the cell count is very low, the PBS volume can be reduced 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 testing. Other biological samples: Centrifuge at 1000 × g for 20 minutes, and remove the supernatant for testing. Sample Appearance: The sample should be clear and transparent, and any suspended matter should be removed by centrifugation. Sample Storage: Samples collected for testing within one week can be stored at 4°C. If testing is not possible, aliquot the sample into single-use aliquots and freeze at -20°C (for testing within one month) or -80°C (for testing within six months). Avoid repeated freeze-thaw cycles. Hemolysis of the sample can affect the final test results, so hemolyzed samples are not suitable for this test. Pre-Assay 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 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: Add 500 μL of universal diluent to each of seven EP tubes. 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 directly as a blank well; there is no need to aspirate the liquid from the penultimate tube, as shown in the figure below. 3. Prepare HRP antibody working solution: 15 minutes before use, centrifuge 100 μg of concentrated HRP antibody at 1000 × g for 1 minute. Dilute the 100 μg of concentrated HRP antibody to a 1 μg working concentration with universal diluent (e.g., 10 μL of concentrated solution + 990 μL of universal diluent). Prepare immediately before use. 4. Preparation of 1× Wash Buffer: Dissolve 10 mL of 20× Wash Buffer in 190 mL of distilled water (Concentrated Wash Buffer removed from the refrigerator may crystallize, which is normal. Allow to stand at room temperature until the crystals have completely dissolved before preparing the buffer). 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 them 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 sealant and incubate at 37°C for 60 minutes. (Recommendation: Dilute the samples to be tested at least 1-fold with Universal Diluent before adding them to the plate. This minimizes the impact of matrix effects on the test results. When calculating sample concentrations, multiply by the corresponding dilution factor. It is recommended to run replicates for all samples and standards.) 3. Wash: Discard the liquid and add 300 μL of 1x wash buffer to each well. Let stand for 1 minute, shake off the wash buffer, and pat dry on absorbent paper. Repeat this process three times (a microplate washer can also be used). 4. Add HRP detection antibody: After washing, add 100 μL of HRP detection antibody working solution directly to each well. Cover with a film sealer and incubate at 37°C for 60 minutes. 5. Wash: Discard the liquid and wash the plate five times according to the procedure in step 3. 6. Add substrate: Add 90 μL of substrate (TMB) to each well, cover with a film sealer, and incubate at 37°C in the dark for 15 minutes. 7. Add 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 450 nm. Calculation of Experimental Results: Result Interpretation: 1. Calculate the average OD value of the standard and sample replicates and subtract the OD value of the blank well as the correction value. 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 it appropriately and retest. Multiply the sample concentration by the corresponding dilution factor when calculating the sample concentration. |
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| Sensitivity | 0.06 ng/mL | ||||||||||||||||||||||||||||||
| Theory | This kit uses a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Sample, standard, and HRP-labeled detection antibody are sequentially added to microwells pre-coated with human DOPA decarboxylase (DDC) capture antibodies. After incubation and washing, the sample is developed with 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 human DOPA decarboxylase (DDC) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | ||||||||||||||||||||||||||||||
| Source | Human | ||||||||||||||||||||||||||||||
| Synonym | Human Dopa Decarboxylase ELISA Kit | ||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | ||||||||||||||||||||||||||||||
| Composition |
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| Background | Aromatic L-amino acid decarboxylase (AADC or AAAD), also known as dopa decarboxylase (DDC), tryptophan decarboxylase, and 5-hydroxytryptophan decarboxylase, is a lytic enzyme. This enzyme uses pyridoxine phosphate (PLP), the active form of vitamin B6, as a cofactor. PLP is essential for the decarboxylation mechanism of AADC. In the active enzyme, PLP binds to lysine-303 of AADC as a Schiff base. | ||||||||||||||||||||||||||||||
| 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, cell lysates and other biological fluids |
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4.2 ★★★★★
Based on 11 reviews
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Product Reviews
★★★★★ 4
Great, except for the edible one
Flavor Name: Chicken & Original, Size: X-Small
My elderly dog and my puppy both love these bones. They are 9lbs and 4.5lbs respectively and this is a good size for them. They also like the next size up and it is manageable for them as well.
The only gripe I have is the edible one, it comes apart in larger chunks (that are still very hard) and my dogs swallowed a few before I noticed how large the pieces were. I decided to throw that one away. Its been over a week and no negative side effects so maybe just me being overly cautious. For the price (gold box) just the other two bones were worth it. They last pretty much forever for my dogs, and I would say they are moderate chewers.
4 stars due to the issue with the edible bone.
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Reviewed in the United States on June 4, 2013
★★★★★ 5
Highly recommend
Flavor Name: Chicken & Original, Size: X-Small
My small dog likes a lot thank you for having i be buying more to give to my family too ❤️ buy you dont want to miss out
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on April 18, 2026
★★★★★ 5
My little dog loves these.
Flavor Name: Bacon & Peanut Butter, Size: X-Small, Flavor Name: Bacon & Peanut Butter, Size: X-Small
These are the perfect size, texture, and density for my 9 lb chihuahua (rescued, of course! :)) He has a tiny mouth with tiny teeth and very little bite force so he needs a chewybone with a lot of give. I've been impressed with how clean these help keep his teeth (it's the saliva flow during the chewing that breaks down any lingering debris and the action of the nubs against the teeth and gums helps to rub off tartar and plaque, if you care to know the *why* behind it.) These are ideal for dogs like mine, and since small dogs tend to have a lot of dental problems due to overcrowded mouths, I couldn't be more grateful that these exist. These would probably also be good for the many rescued puppy mill dogs, who are often small breeds, who have dental issues due to malnutrition and exacerbated by overbreeding, especially in females whose bodies often rob the jaw bones of calcium for the sake of the fetuses they carry, when the pregnant dogs aren't given an appropriately enriched gestational diet. That leads to the little bleppy tongues sticking out all over that we often see in the canine survivors of back yard breeding operations, puppy mills, and the like. There are a lot of these dogs being rescued and adopted in the U.S. right now and these would likely be ideal for them. (In case any adopters of these little survivors are reading this. :)) These are for gentle chewers only, and there are a lot of dogs who fall into that category. They aren't , however, for dogs with an average bite or higher, which is where some of the complaints you hear about these are coming from. Know your dog's chewing needs and buy the size that fits their mouth, tooth type, and bite strength. My dog barely puts a dent in these after a year or more of chewing (watch for any degradation of the plastic and replace as needed, of course.) For little dogs like mine, these are just fantastic and I hope Nylabone keeps making these type of chew toys to keep our dogs healthy. :)
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Reviewed in the United States on February 22, 2019
★★★★★ 3
he only like the Dark one
Flavor Name: Bacon & Beef, Size: X-Small
love nylabones for my pup
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Reviewed in the United States on April 29, 2026
★★★★★ 5
My Dog's Favorite Nylabone
Flavor Name: Bacon & Beef, Size: X-Small
I've been testing out Nylabones the last couple months for my chihuahua puppy. He's a heavy chewer and too easily tears apart many of them, then others are too hard and I take them away because I fear for his teeth. This set is his favorite so far. He doesn't care about the peanut butter ones, loves the beef one, and is absolutely obsessed with the bacon one. He walks around with it in his mouth whenever he goes anywhere. Unfortunately, my senior chihuahua also loves the bacon one and there is now an ongoing feud over it, so guess I'm buying another pack. I like that these are just a little chewy without being too soft, but I still recommend keeping an eye on them as even my puppy has managed to start to get little pieces off. But it's more as frayed strings than chunk like I've had happen with some of the softer ones.
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Reviewed in the United States on February 22, 2023