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Description
Mouse Nestin ELISA KitProduct Specification Usage Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High precision pipette and gun 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: Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4 overnight, then centrifuge at 1000g for 20
Product Specification
| Usage |
Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High-precision pipette and gun 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: Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4℃ overnight, then centrifuge at 1000×g for 20 minutes, and take the supernatant, or store the supernatant at -20℃ or -80℃, but avoid repeated freezing and thawing. Plasma: Collect the specimen using EDTA or heparin as an anticoagulant and centrifuge at 1000 × g for 15 minutes at 2-8°C within 30 minutes of collection. The supernatant can be assayed or stored at -20°C or -80°C, but avoid repeated freezing and thawing. Cell Supernatant: 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., 1 g of tissue sample to 9 mL 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 the tissue cells, the homogenate can be sonicated or repeatedly frozen and thawed. Finally, centrifuge the homogenate at 5000 × g for 5-10 minutes, and the supernatant can be assayed. Preparation before testing: 1. Please remove the test kit from the refrigerator 10 minutes in advance and equilibrate to room temperature. 2. Preparation of gradient standard working solution: Add 1mL of universal diluent to the lyophilized standard, let it stand for 15 minutes to completely dissolve, and then gently mix (concentration is 10ng/mL). Then dilute to the following concentrations: 10ng/mL, 5ng/mL, 2.5ng/mL, 1.25ng/mL, 0.625ng/mL, 0.3125ng/mL, 0.15625ng/mL, and 0ng/mL. Serial dilution method: Take 7 EP tubes and add 500uL of universal diluent to each tube. 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 step to mix thoroughly. 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 Nestin capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB converts to blue under the catalysis of HRP and to yellow under the action of acid. The intensity of the color is positively correlated with the amount of Nestin in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Mouse | |||||||||||||||||||||||||||||||||
| Synonym | Mouse Nestin ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Nestin is a protein encoded by the NES gene. Nestin is a type VI intermediate filament (IF) protein. These IF proteins are primarily expressed in neurons and are involved in radial axonal growth. Seven genes encode the heavy neurofilament (NF-H), medium neurofilament (NF-M), and light neurofilament (NF-L) proteins: nestin and α-internexin in neurons, synemin α and desmuslin/synemin β (two alternative transcripts of the DMN gene) in muscle cells, and syncoilin (also in muscle cells). Most members of this group prefer to associate in tissues as heteropolymers. Nestin forms homodimers and homotetramers in vitro but does not form IFs on its own. In mixtures, nestin preferentially co-assembles with purified vimentin or the type IV IF protein internexin to form heteromeric coiled molecules. | |||||||||||||||||||||||||||||||||
| 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, cell supernatant, tissue homogenate, etc. |
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4.4 ★★★★★
Based on 25 reviews
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Product Reviews
★★★★★ 3
Not for chewers
Flavor Name: Bacon & Chicken, Size: Small
My dog liked these okay. He was able to chew little pieces off though.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on March 29, 2026
★★★★★ 5
Great fit
Flavor Name: Peanut Butter, Size: X-Large
My dog has a very powerful jaw. Only a few things that he cannot destroy.
We have had the peanut butter flavor that still looks the same that it did a year and a half ago. It is his version of a binky. He may leave it alone for days or weeks but every so often he needs to have his teeth and gums itched.
Very few remnants of the bone left around.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on June 10, 2026
★★★★★ 5
A Humorous Twist on Tough Dog Toys!
Size: Large, Color: Lobster 3 Pack
I recently purchased the Nylabone Power Chew Dog Toy Pack for Aggressive Chewers from Amazon, and I must say, these toys have been a hilarious hit with my furry friend. As a dog parent of a strong and determined chewer, finding toys that can withstand their chewing prowess is a top priority, and these toys deliver both durability and a touch of humor.
The pack consists of three large/giant-sized toys in the shape of a lobster, cheese, and broccoli. The unique and amusing design puts a smile on my face every time I see my dog interacting with them. The unexpected twist brings a playful element to our daily playtime sessions, making it a fun and engaging experience for both of us.
When it comes to durability, these toys excel. My dog can be quite aggressive with their chewing, and I've struggled to find toys that can withstand their powerful jaws. However, the Nylabone Power Chew toys have proven to be impressively tough and durable. Despite rigorous chew sessions, these toys have remained intact, with no signs of wear or damage. They provide a reliable outlet for my dog's chewing instincts without the risk of shredding or ingesting any harmful materials.
The textured surfaces of these toys add an extra layer of interest for my dog. The ridges and raised bumps provide sensory stimulation, promoting healthy oral hygiene and helping to clean teeth and gums as they chew. I appreciate the added dental benefits these toys offer, as it helps maintain my dog's dental health while they engage in play.
I also appreciate the size options available in this pack. Large and giant-sized dogs often require bigger toys to accommodate their larger mouths and chewing strength. The included toys are appropriately sized for larger breeds, ensuring a comfortable grip and maximum enjoyment during play sessions.
As a bonus, these toys are made from high-quality, non-toxic materials, so I have peace of mind knowing that my dog can safely interact with them. Safety is a top priority for me, and Nylabone consistently delivers in that regard.
In conclusion, if you have an aggressive chewer and are in search of durable, funny, and engaging dog toys, the Nylabone Power Chew Dog Toy Pack is a fantastic choice. These toys provide hours of entertaining play, withstand even the most determined chewers, and offer dental benefits. Embrace the humorous twist and enjoy watching your furry friend engage in safe and engaging play with these fantastic toys.
Invest in the Nylabone Power Chew Dog Toy Pack today and let the lobsters, cheese, and broccoli bring laughter and durability to your dog's playtime!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on December 19, 2023
★★★★★ 5
Solid win for any chewing pup!
Size: Small, Color: Broccoli 3 Pack, Size: Small, Color: Broccoli 3 Pack
My pup loves them! Long lasting plus health benefits for his teeth, 10/10 would recommend 💖
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on April 27, 2026
★★★★★ 5
Shark dog approved
Size: Large, Color: Broccoli, Size: Large, Color: Broccoli
My new baby shark.... I mean puppy really enjoys chewing. I can't just give her a ton of pig ears or bully sticks all day, which she would definitely love, so I'm constantly looking for well made chew toys. I'd put this right up with Kong. Very durable and nice. I also love the different textures, she seems to also love it. She can gnaw at the top and chew on the stem. She wanted it as soon as I opened it - it does smell like bacon but I haven't taken a chomp at it to be able to write a complete review!
My dog is about 23 lbs but I can see a 80 lb dog chew this, it's not small.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on October 8, 2025
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