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Description
Mouse NNMT 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. Centrifuge the specimen 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. 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 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., 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 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. 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 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 500uL of universal diluent to each. 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 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 nicotinamide-N-Methyltransferase (NNMT) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by HRP peroxidase and to yellow by acid. The intensity of the color is positively correlated with the amount of nicotinamide-N-Methyltransferase (NNMT) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Mouse | |||||||||||||||||||||||||||||||||
| Synonym | Mouse Nicotinamide-N-Methyltransferase ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Nicotinamide-N-methyltransferase, also known as NNMT, is an enzyme encoded by the NNMT gene. It catalyzes the methylation of nicotinamide and similar compounds using the methyl donor S-adenosylmethionine (SAM-e) to produce S-adenosyl-L-homocysteine (SAH) and 1-methylnicotinamide. Nicotinamide methylation by NNMT and SAM-e is the primary pathway of nicotinamide degradation leading to urinary excretion. It is highly expressed in the human liver. Its expression in human embryonic stem cells is thought to help maintain cells in a naive state. NNMT expression is significantly upregulated in many cancers, including pancreatic cancer, where elevated NNMT enzyme levels are associated with an increased risk of mortality. | |||||||||||||||||||||||||||||||||
| 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 | Serum, plasma, tissue homogenates and other biological fluids |
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4.2 ★★★★★
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Product Reviews
★★★★★ 5
Nice quality for a good price
Color: Luminous watch black face
Bought this for my boyfriend for Christmas. He loves it! Nice coloring, looks like the pictures. Fits nicely and comfortably once the size was adjusted. He did have some difficulty in sizing it - the little tool provided could’ve been of better quality to make this process easier but it did the job.
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Reviewed in the United States on March 9, 2026
★★★★★ 3
3/5 Stars – Looks far more expensive than it is
Color: Luminous watch black face, Color: Luminous watch black face
The OLEVS 5885 is about what you’d expect from a ~$30 watch, but in a good way. I bought it knowing it wasn’t going to compete with higher-end brands, and with that expectation in mind I’ve actually been pretty impressed.
The biggest standout is the appearance. This watch looks far nicer than its price point would suggest. I get compliments on it fairly regularly from both friends and random people who assume it cost much more than it did. If you’re looking for something stylish on a budget, it definitely delivers in that department.
Functionally, it does its job well enough. It keeps time reliably and hasn’t given me any real issues there. The glass/crystal is also surprisingly durable. I’ve worn this watch quite a bit, and while the rest of the case and bezel have picked up scratches over time, the face itself has held up extremely well.
Another pleasant surprise is the bezel. It’s functional and unidirectional, with strong, clear, tactile clicks. It feels solid and hasn’t loosened up with use.
That said, the cheaper elements do show up if you look closely. The bezel markings are pretty cheap and scratch easily. Also, the small lume marker at the 12:00 position on the bezel fell out during the second week of owning the watch.
Overall though, I’m quite happy with it. For the price, it looks much more high-end than it really is, and I can comfortably use it as an everyday tool watch without worrying about babying it like a luxury piece.
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Reviewed in the United States on March 12, 2026
★★★★★ 5
Good cheap watch. Watch band could be better
Color: Luminous watch black face
The timepiece itself is use of decent quality and is perfectly acceptable for everyday use. I'm pretty rough on my watches and this watch fits the bill as I go about my daily routine. It keeps good time and has a good looking face. It seems many have trouble with the watch band sizing. The arrows on the inside of the watch band show the direction the pins should be pushed out. The pins stick out ever so slightly on one side or the other of the watch band giving the band a rough feel. I used a soft nail file and polished the pins smooth. The watch band feels much better afterwards. I would buy it again and recommend the watch to anyone looking for a cheap timepiece.
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Reviewed in the United States on January 21, 2025
★★★★★ 4
OLEVS divers watch in stainless steel
Color: Luminous watch black face
Never owned a divers watch and especially not one
by Olevs. Glad I bought it great quality and price
looks great too.
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Reviewed in the United States on March 5, 2026
★★★★★ 1
unless you are adept at figuring out how to fix things , don't buy.
Color: Luminous watch black face
This watch looks good but it's not easy to remove the links , to make it shorter. Also the clasp broke right away! It was a waste of money for me.
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Reviewed in the United States on March 25, 2025