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Description
Rat SIRT1 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: Gently wash adherent cells with pre-chilled PBS, then trypsinize and collect the cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be collected 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 (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. Cell Culture Supernatant: Centrifuge at 1000×g for 20 minutes. Collect the supernatant for analysis, or store at -20°C or -80°C, but avoid repeated freezing and thawing. 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 enzyme conjugate are added sequentially to microwells pre-coated with a Sirtuin 1 (SIRT1) capture antibody. 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 Sirtuin 1 (SIRT1) in the sample. The absorbance (OD value) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Sirtuin 1 ELISA kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Sirtuin 1, also known as SIRT1 or NAD-dependent deacetylase sirtuin-1, is a protein encoded by the SIRT1 gene. It is an enzyme primarily located in the cell nucleus. It is a member of the sirtuin protein family and is a homolog of the Sir2 gene in Saccharomyces cerevisiae. Members of the sirtuin family are characterized by a sirtuin core domain and are divided into four classes. The function of human sirtuins has not yet been determined; however, yeast sirtuins are known to regulate epigenetic gene silencing and inhibit rDNA recombination. Research suggests that human sirtuins may function as intracellular regulatory proteins with mono-ADP-ribosyltransferase activity. The protein encoded by this gene belongs to class I of the sirtuin family. It is downregulated in cells with high insulin resistance, and inducing its expression increases insulin sensitivity, suggesting that this molecule is involved in improving insulin sensitivity. Furthermore, it has been shown to deacetylate and influence the activity of two members of the PGC1-α/ERR-α complex, essential metabolic regulatory transcription factors. In mammals, it has been shown to deacetylate, thereby inactivating the p53 protein. It also stimulates autophagy by preventing the acetylation (via deacetylation) of proteins required for autophagy, as demonstrated in cultured cells, embryonic, and neonatal tissues. This function provides a link between sirtuin expression and the cellular response to limited nutrients due to caloric restriction. It plays a role in activating T helper 17 cells, which contributes to autoimmune diseases. Efforts to therapeutically activate SIRT1 may trigger or exacerbate autoimmune diseases. Together with HDAC1 and the AP-1 promoter complex in D1-type dopaminergic spiny neurons, it appears to be closely implicated in the pathogenesis of addiction. | |||||||||||||||||||||||||||||||||
| 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, cell culture supernatants, and other biological fluids |
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4.1 ★★★★★
Based on 25 reviews
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Product Reviews
★★★★★ 5
Easy and simple: Exact Step-by-Step Guide to Reloading Your Balance
Reload type: One Time Reload
I found reloading my Amazon gift card balance much easier than expected. The process was simple and took about 5 minutes at most. Here are the exact steps to do it:
Login: In the Amazon app or on Amazon.com, log in to your account.
Menu: Tap the 3 lines on the bottom right corner of your screen (next to Rufus, Amazon's AI).
Find Category: In the section highlighted "Shop by category," tap on Gifting & Registry (3rd option down in that section) and a drop-down menu will appear.
Select Gift Cards: On the drop-down menu, tap Gift Cards (first option).
Choose Your Action: "The Gift Card Shop" will appear. There you will have three options: "Redeem a gift card," "View your balance," and "Reload."
Start Reload: To add funds, tap Reload.
Select Type: Scroll down to "Add funds directly to your Amazon Gift Card balance" and tap which reload type you want (one-time, low balance auto-reload, or scheduled auto-reload).
Enter Amount: Choose your reload amount. The default is $25, but you can type in your own amount as well (the minimum amount is $5.00).
Finish: Tap the orange "Buy Now" tab.
***I used the Amazon app using the u i OneUI version 8.0 these Amazon app on Android version 16 (Baklava) on a Samsung Galaxy S25 Ultra. Others may have a slight variation of the menu setup on the Amazon app.***
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on April 22, 2026
★★★★★ 5
Easy way to add balance
Reload type: One Time Reload
Reloading my Amazon gift card balance has been easy and convenient. I like using it when I want to keep a little money set aside for Amazon orders or make checkout quicker without thinking about which card I want to use. The process is straightforward, the balance shows up fast, and it has been reliable every time I have used it. It is also helpful for budgeting because I can add a set amount and then use that balance for regular purchases. Nothing complicated about it, which is exactly why I like it.
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Reviewed in the United States on May 28, 2026
★★★★★ 5
Monthly reload for Subscribe & Save Orders
Reload type: One Time Reload
Not a glamorous review, but a genuinely useful tip for Subscribe & Save users. I reload about $100 to my gift card balance once a month, a week or two before my S&S order processes. Amazon pulls each Subscribe & Save item as a separate transaction, which means 10+ individual charges hitting your bank account on before or on ship day. A headache for budgeting and a minor anxiety trigger if you're watching your account for unauthorized activity. Loading the balance first consolidates everything into one clean transaction. Your S&S order then pulls from the gift card balance rather than your card directly, and your bank statement stays tidy.
The side benefit is built-in budget discipline. I set about $100 as my monthly Amazon ceiling - if my cart runs over, something gets pushed to next month. It's a simple system that keeps spending intentional without any extra effort. If you're a regular S&S user who wants cleaner finances and less ""wait, who charged this and what was that charge?"" moments, this is worth making a habit.
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Reviewed in the United States on June 6, 2026
★★★★★ 5
Great for Budgeting – Instant Balance and Easy to Use
Reload type: One Time Reload
I use Amazon’s gift card reload feature to plan my spending. It’s perfect when I know I want to make a purchase but prefer to set the funds aside first.
I love that it’s immediate—no worries about pending charges or accidental spending from my card. It gives me peace of mind knowing that when I’m ready to buy, the balance is just sitting there, waiting.
Pros: easy to use, instant balance update, helps with budgeting and avoiding overspending
Cons: funds are locked into Amazon once reloaded, doesn’t count toward certain account requirements like review eligibility
Overall, it’s a convenient and reliable way to manage spending on Amazon, especially if you like having control over your purchases.
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Reviewed in the United States on May 1, 2026
★★★★★ 5
I did this with a pay in 4. It made it super simple for a higher balance.
Reload type: One Time Reload
I’ve used this to reload my balance a few times, and it’s been simple and reliable. The process is quick, and the funds show up without any delays or issues.
It’s a convenient option when you want to control spending or keep purchases separate without using a card each time. Everything is straightforward and easy to manage.
I haven’t run into any problems with transactions or balance updates. It’s been consistent every time I’ve used it.
Overall, it’s an easy and dependable way to keep funds ready to go without any hassle.
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Reviewed in the United States on May 6, 2026