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Description
Human RTN4R 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 utilizes 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 Reticulon 4 Receptor (RTN4R) 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 Reticulon 4 Receptor (RTN4R) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Reticulon 4 Receptor ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Reticulin 4 receptor (RTN4R), also known as Nogo-66 receptor (NgR) or Nogo receptor 1, is a protein encoded by the RTN4R gene. This gene encodes a receptor for reticulon 4, oligodendrocytemyelin glycoprotein, and myelin-associated glycoprotein. This receptor mediates axonal growth inhibition and may play a role in regulating axonal regeneration and plasticity in the adult central nervous system. The Nogo-66 receptor (NgR) is a high-affinity binding receptor for a region of Nogo, a myelin-associated protein that inhibits axonal growth. | |||||||||||||||||||||||||||||||||
| 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 | 0.31-20 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, and other biological fluids |
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4.5 ★★★★★
Based on 90 reviews
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Product Reviews
★★★★★ 1
Not as safe as it seems
Size: Xtra Large
I did like the look and sound of the all natural wood and coconut rope. However after 15 minutes of giving it to my dog there were rope fibers everywhere. Even though my research said coconut rope was ok for dogs in small amounts; my dog started throwing up. My cat also managed to get ahold of some of the strings and ended up choking. Even when I took the rope off and gave just the coffee wood stick; after just a few minutes of play my dog began coughing. This is just the experience that I had. All dogs are different. Just please don’t let your best friend play with it unsupervised
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on February 3, 2025
★★★★★ 5
Best chew toy for a small dog
My Pomchi puppy was chewing on anything and everything, including me. He had a million other toys that he just didn’t care about. This is small and perfect for his tiny, ferocious mouth. It seems very durable so far and believe me, he doesn’t go easy on this thing. It’s hard enough that I can tell it feels good on his tiny, razor sharp teeth. The bone keeps him busy all throughout the day, and it doesn’t squeak, which is good because squeaky toys freak him out. I highly recommend this if you have a tiny, teething monster.
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Reviewed in the United States on September 27, 2025
★★★★★ 5
Pomeranian perfect size
Small size perfect Pomeranian (1 yr old) she prefers over other chews - have ordered twice
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Reviewed in the United States on April 10, 2026
★★★★★ 5
My puppies favorite bone to chew
I’ve bought at least a half a dozen of puppy bones for my doggie to try, the majority being from Nylabone, and the others from hartz. Her first bone was a pink puppy Nylabone xsmall sise. She definitely must have liked that one but unfortunately I felt like I had to take it away from her too soon, she was teething more then, but even so, thought it was unsafe when I saw little pieces come off and her trying to chew them.
I really like these the best, though I don’t think I’ll be ordering any more at least not until she finishes her new ones. I don’t know what flavor she likes more, it might even be a color thing, but I think she prefers the orange ones which are peanut butter flavored. These are really ideal for toy breeds. My pup is still growing, she’s almost 7 months now, but unsure when she will stop growing, for reference she is a chocolate sable yorkiepoo, not sure of how much she weighs right now, but would estimate she weighs around 5.5-6 pounds. These are perfect for her, and I feel like they out live the other ones I’ve gotten, or she just has more interest in them. I bought a second package on subscribe and save (or possibly on my first and forgot I had a new order and didn’t return), but didn’t open the new package until recently as I felt the first pack were starting to look a bit knawed off, but in the middle there was still plenty of chew left. Oddly, she has prefered the old orange one vs the brand new orange one despite them being the exact same toy and flavor. I think she just hasn’t broken it in yet and I overloaded her with too many toys! I also got them for a great price, for some reason I noticed that the price increased nearly double of what I paid, and now they habe decreased a bit but still are about $2 more than what I paid. She loves all toys, and has many plush kinds. She still teeths, so when she is extra hyper and such, I feel like the bone helps occupy her for a moment. All in all I’d reccomend these out of any puppy bones to chew for toy breeds!
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Reviewed in the United States on April 2, 2022
★★★★★ 4
Nice chew toy
Puppy likes but for me to hold so she can chew is a little difficult.
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Reviewed in the United States on May 8, 2026
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