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Description
Human Klotho 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: 1. 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: 1. 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. 2. 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. 3. 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. 4. 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 enzyme conjugate are sequentially added to microwells pre-coated with a Klotho capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by HRP peroxidase and then to yellow by acid. The intensity of the color is positively correlated with the Klotho content in the sample. Absorbance (OD) is measured at 450 nm using a microplate reader to calculate sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Klotho ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Klotho is a transmembrane protein that regulates insulin sensitivity. It possesses β-glucuronidase activity, hydrolyzing β-glucuronic acid and is associated with human aging. Circulating klotho levels in plasma decrease with age. β-Klotho acts as a co-receptor with FGF receptors and can bind to fibroblast growth factors (FGFs) such as FGF19, FGF20, and FGF23. The mechanism of action of klotho remains unclear, but it is known to regulate cellular calcium homeostasis by increasing TRPV5 expression and decreasing TRPC6 expression. Klotho also increases expression of the inwardly rectifying ion channel ROMK. | |||||||||||||||||||||||||||||||||
| 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, tissue homogenates and other biological fluids |
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4.7 ★★★★★
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Product Reviews
★★★★★ 1
good ball, wrong color
Size: Medium
I think the hot-ee roller is a great ball! My problem is that there is no option to select a color. I thought I was getting a purple one, since that is what is shown in the picture. As usual, I failed to read that it was assorted colors. I have a peculiar little dog that will only play with the purple one. His current purple one is in tatters. You could hardly even call it a ball at this point. We have red, blue and green and they are invisible to him.
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Reviewed in the United States on May 14, 2026
★★★★★ 5
Makes my melanated skin POP!
Wow! Where has this product has been all of my life??? Up until now, I believed that dewey, glowing, buttery skin I saw on celebrities in photos and video were due to some ultra expensive product that I could not afford, but low and behold I found out that is NOT true and you can achieve that same look if you're "balling on a budget", like ME! While Vaseline, and baby oil are mainstays, sometimes you need something with a little bit more umph, sheen, and sparkle and THIS Vaseline oil does the trick and then some! One mistake I made when I first started wearing it: I didn't spend enough time rubbing it into my skin. If you don't take the time to really rub it in and allow your skin to soak it in, you will have some dry spots (i.e., areas of skin you thought you moisturized but actually missed). I now understand why makeup artists and stylists are always seen rubbing their clients legs and arms, and between pictures and takes- it's because they're making sure they get the product into the skin! That said, this is not something that you can just slap on and go. You have to really take your time and be intentional about the application. If you have a partner, roommate, spouse, or whatever, ask them to help rub it into your back. I also love putting this on after I get out of the shower and I want to wear something sexy to bed-It is a major confidence booster when you are wearing lingerie. I absolutely LOVE this product and it is the best thing I ever bought for my skin. I bought both golden hour glow and sunlit glow. The sun lit is good for a lightly glazed look but the golden hour glow is a lot more sparkly and has this gold, copper, Sheen to it - basically, it's heavier or more pigmented, but still very lightweight. It does not stain in my clothes, it is very soft and it gives my skin this glazed, sun kissed, glow that is absolutely out of this world! The oil quality is great because it's not too thick and it's not too thin and it doesn't need to be reapplied. There is virtually no scent, so it pairs well with perfume or body spray. I have a very sensitive nose, so I was a little bit concerned about a strong fragrance but it does not have that at all. It is so versatile: you can use it on feet, legs, back, chest, forearms--and a little goes a long way. This is absolutely amazing. It is much better than the previous Vaseline bronze oil that I wore that was a lot more watery and came in a pump- it just didn't do it for me. This line is more moisturizing - this glazed and glisten gel is the best thing ever! Run, do not walk, to buy this product--You won't be disappointed. It is great value for the money. 10 stars!!!!
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Reviewed in the United States on May 23, 2026
★★★★★ 5
Really loving this!
This gives such a pretty sun-kissed glow without looking orange or over-the-top glittery. I’ve been using it on my arms and legs and it makes my skin look healthy, hydrated, and glowy without feeling greasy. It’s lightweight, easy to apply, and lasts longer than I expected. Smells good too. Definitely feels like a great value for the money. Perfect for summer, vacations, or when you just want a little extra glow without self tanner.
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Reviewed in the United States on May 17, 2026
★★★★★ 5
Beautiful glow
The way this makes my skin looks is heavenly. I’ve been using it on vacation and it makes my skin feel so smooth. The product is lightweight and doesn’t feel sticky on skin and the shimmer lasts all day. There’s no strong smell which I’m super happy about. Would definitely recommend this.
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Reviewed in the United States on May 27, 2026
★★★★★ 4
Does not stain! Great body oil
It’s soooo light weight and easy to apply, you don’t need tons of it to get a nice result. It’s not very oily nor greasy at all. It DOES NOT STAIN!!!! I love that, it moisturized my skin so well, it looks so pretty and natural but I’m going to mention that the scent is a little weird, not bad but like I was expecting to smell like vanilla but it’s not vanilla… it’s something else but is ok because it’s not strong or unpleasant.
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Reviewed in the United States on May 27, 2026