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Description
Human NELL2 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 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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| 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 protein kinase C-binding protein NELL2 (NELL2). 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 protein kinase C-binding protein NELL2 (NELL2) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Protein kinase C-binding protein NELL2 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | NEL-like protein 2, also known as NELL2, is an enzyme encoded by the NELL2 gene. This gene encodes a cytoplasmic protein containing epidermal growth factor (EGF)-like repeats. The encoded heterotrimeric protein may be involved in cell growth regulation and differentiation. Several variants exist, encoding several different isoforms, at least one of which appears to be a secreted protein. Research suggests that this protein plays a role in neuronal cell growth and differentiation, as well as tumorigenesis. It is involved in regulating hypothalamic GNRH secretion and controlling puberty. | |||||||||||||||||||||||||||||||||
| 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, and other biological fluids |
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4.6 ★★★★★
Based on 14 reviews
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Product Reviews
★★★★★ 5
Nylabone Ring Bone
Size: X-Small, Color: Ring Bone
My dog loves these. She chews them into pieces which helps keep her teeth clean. It is a mess to clean up. She chews and spits out the pieces. She is 6 years old so not a puppy.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on May 29, 2026
★★★★★ 4
Everything a Teething Puppy Could Need — Well-Thought-Out Kit
Size: X-Small, Color: Teething Bone
The Nylabone New Puppy Starter Kit is a solid all-in-one solution for the first few months of puppy ownership. It comes with multiple chew toys and bones designed specifically for teething puppies, and my little one took to them right away. The variety keeps him engaged, and I appreciate that each item is designed to be gentle on developing teeth while still satisfying his urge to chew.
What differentiates this kit from other puppy chew sets on Amazon is the thoughtful selection of textures and shapes. It’s clear these toys are designed to stimulate gums, promote healthy chewing habits, and even support dental care. I also like that everything is non-toxic and durable enough to survive a few enthusiastic chews.
The only reason I didn’t give 5 stars is that the kit might be a bit small for larger breeds, or for puppies that are particularly aggressive chewers. But for standard small to medium pups, it’s a fantastic starter kit that keeps both puppy and owner happy.
✅ Bottom line: A reliable, well-designed starter kit for teething puppies which is thoughtful, durable, and engaging. Makes a great gift for new puppy owners or a practical addition to your own puppy’s toy collection.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on September 26, 2025
★★★★★ 5
Chew
Size: X-Small, Color: Ring Bone
My dog loves this little chewy bone and it’s good for her teeth. That was very cheap on a sale. It’s worth giving it a try highly recommend it.
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Reviewed in the United States on June 11, 2026
★★★★★ 3
Chicken Flavor Good, But 🔄 Rotation-Resistant✖; Best for Simpler Chewers
Size: X-Small, Color: Ring Bone, Size: X-Small, Color: Ring Bone
🐾 Hi, I’m PVD; plush sophisticate, chew critic, and active curator of rotational toy relevance.
This ring bone and I never fully bonded. I sampled it maybe five times in my entire puppyhood before quietly returning it to the bin (with a single judgmental glance).
Still, it does have value for the right pup.
⭐ Why I originally wanted to love it:
✔️ XS size works well for very young puppies
✔️ Chicken flavor lasts impressively long
✔️ Freezing offers temporary gum relief
✔️ Durable and safe; no odd odors or shredding
🌀 Why it’s now a respectful 3-star chew in my rotation:
✘ Never became a go-to toy (5 uses max in my life)
✘ Ring shape felt awkward, not natural or chic; hard to grip and maneuver
✘ Texture too tame vs. nubs or detailed chew surfaces
✘ When I was smaller, I much preferred the Nylabone Baby Blue Bone Chew Toy with nubs (my personal classic), and have since matured into nightly Nylabone “Nubz” edible bones
This Ring Bone is Best For:
• Very young puppies with minimal chew preferences
• Dogs who like simple textures and circular shapes
• Those just starting enrichment chew play
• Freezer treat rotations (briefly)
🚫 Not ideal for: picky pups, advanced chewers, or dogs who analyze toy ergonomics (like myself).
📝 Final Re-Barks:
A strong, well-made chew, but lacked the intrigue to make my permanent rotation. Works for basic chew enjoyment; not for pups with elevated taste or toy ambition.
— PVD (Chew Strategist, Textural Analyst & Aspire-to-Vine Reviewer 🐶💼)
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Reviewed in the United States on September 17, 2025
★★★★★ 5
Puppy loves it!
Size: X-Large, Color: Teething Bone, Size: X-Large, Color: Teething Bone
My small puppy loves this bone! I originally purchased this for a larger dog, but my small 10 pound puppy loves it more. It’s good quality and a softer material for puppies!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on May 10, 2026