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Description
Rat RIPK3 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 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 receptor-interacting serine/threonine-protein kinase 3 (RIPK3) 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 receptor-interacting serine/threonine-protein kinase 3 (RIPK3) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Receptor-interacting serine/threonine-protein kinase 3 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Rat RIP3 factor receptor-interacting protein 3 (RIPK3), also known as receptor-interacting serine/threonine protein kinase 3, is an enzyme encoded by the RIPK3 gene. The gene product is a member of the receptor-interacting protein (RIP) family of serine/threonine protein kinases and contains a C-terminal domain distinct from other RIP family members. The encoded protein is primarily localized in the cytoplasm and can undergo nuclear-cytoplasmic shuttling in response to novel nuclear localization and export signals. It is a component of the tumor necrosis factor receptor-I signaling complex and can induce necroptosis through interaction with RIPK1 and MLKL in a protein complex known as the necrosome. | |||||||||||||||||||||||||||||||||
| 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 | Tissue homogenates, cell lysates, and other biological fluids |
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4.6 ★★★★★
Based on 8 reviews
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Product Reviews
★★★★★ 4
Very good watch
Color: White/Chrome-tone
Was a gift for my girlfriend. I even liked it has a mode for two different times sets.
The strap does get dirty fast. But its really useful this watch and very good in estetics.
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Reviewed in the United States on October 24, 2025
★★★★★ 5
Very nice feminine quality watch
Color: Pink/Rose Gold-Tone, Color: Pink/Rose Gold-Tone
I purchased this watch for work. As a nurse, I need the exact date and time in military time and seconds, so this watch is perfect. It’s the 3rd of this kind that I’ve ordered over the years and this one is definitely my favorite. Much easier to program than previous models and the look is very feminine and cute. Of course being water resistant is important since I have to wash my hands a million times a day. I’m hoping it will last a long time as previous models have. Usually the plastic band breaks from daily wear and tear before the battery has ever stopped, but that’s after years of daily use. That’s why I prefer to stick with this brand. They’ve always been very durable, lightweight, and effective for what I need it for. I would definitely recommend. This version is the cutest, like I would wear it even when not working. So I’m very happy with it.
*** Update:
2 years later I still wear this watch everyday, at work or not. Has never broken, battery still going strong and still so pretty. Awesome 👏🏻
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Reviewed in the United States on June 13, 2024
★★★★★ 5
Great modest, minimalist digital watch: 2 time zones, Indiglo, subtle hourly chime
Color: Purple/Silver-Tone
Great value and this is nice, low-key but functional and sleek timepiece. I bought it to replace an older Ironman (bulkier) watch that finally reached end of life after LOTS of adventures (still functions, but strap and face just really couldn't be taken into corporate or social settings without expectation of some heckling). I REALLY like most of the new features (especially the dual time zones, hourly chime (can toggle on/off), and Indiglo applied to the numbers/text rather than as background), I probably would have switched to this earlier if I had known of them. Slight negative that the hourly chime toggle is nested within the time setting workflow, so can't just nearly instantly switch it when heading into movie/performance/other setting where want to silence it fully. I can't speak to the water resistance of this one yet, have been taking it off to shower and haven't been swimming yet. (Taking it off to shower in part because had seen other reviews noting that the trim around the dial may not be long-lasting. So out of caution to extend its appearance, I'll probably remove it for salt/chlorinated water swims (not timing laps)). Only other aspect should note is that the feature that helps to "lock" the strap band at end (bit of a nub to catch) is bit tricky when putting it on / taking it off, may be even more so for those with arthritis or other condition that would make it extra challenging/frustrating to maneuver and engage.
Quite happy with it overall though, just a slight annoyance when putting on / taking off (which may not apply for most if they just leave it on).
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Reviewed in the United States on January 17, 2023
★★★★★ 3
Good Everyday Watch - But Scratches Easily!
Color: White/Chrome-tone, Color: White/Chrome-tone
This is exactly the watch I was looking for. I needed a basic watch that is waterproof, can also act as a stopwatch, I don’t need to charge, and hold up to regular wear and kid activities. Set up was incredibly easy and didn’t need any directions. The size and style are slim and sleek making it easy to wear under just about anything or wear to bed. I’ve had Iron Man watches in the past that I’ve loved. I lost my recent one hence this replacement. I’m hopeful that this watch can hold up to every day wear and not have any of the wristband or chipping issues as other users have seen. Will come back to update the review after a few weeks of wear.
3/19/26: Back for an update. Watch function is still great but the face of the watch gets scratched so easily. I questioned these reviews when I ordered the watch thinking, “how bad could it be?” And they’re right. Super easily scratched. I have no idea how I even scratched/damaged mine. Doesn’t affect function but frustrated that it happened within a month of use/wear. I suppose you get what you pay for - very disappointing for well known brand such as Timex. Lost two stars for this sole reason.
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Reviewed in the United States on February 12, 2026
★★★★★ 3
Functionally not bad, but style is meh
Color: Blue/Rose Gold-Tone, Color: Blue/Rose Gold-Tone
Likes:
Very easy to read with the large numbers/characters
The “indiglo” lighting seems to efficient in dark/dim light
Dislikes:
Setup is not easy. It did not come with instructions.
Watch time lags behind
Find the watch a little uncomfortable on the wrist
I need a watch for the clinical portion of my schooling. This watch is not terrible. The display is easy to read and the indiglo light displays well. Ultimately, I’ve decided to return it. I wish it had instructions. I got by enough to set the correct time and date, but it appears there are two times you can preset it to. So maybe you can have one set as one time zone & the other another time zone. I was going to set one as normal time and the other as military time, but couldn’t figure out how to set which preset you want to currently display, so I just set both as the same exact thing.
I like a different watch, also purchased from Amazon, better for myself after comparing it to this timex. I just don’t care for the look of this one. It looks a bit too juvenile. My sister described it well as a watch she “would have worn as late elementary/middle schooler” and I agree. I have a feeling that maybe it doesn’t come across as such in the white. White was my first choice, but other reviews mentioned the band dirtying and not being able to remove stains/coloring, so I went with this blue band color. Also, something about the watch was a bit uncomfortable while wearing it and I have a hunch settling in to wearing it more would not solve that discomfort much, if at all. Also, I initially set the watch 2 minutes early against the time on my phone. But it’s now days later and I noticed it runs behind the time on my phone by about 30 seconds. Both the watch I compared to this one and this timex itself claim to be water resistant, but I have not tested those claims with any hand washing or anything. I trust they’ll be fine in that sense. Also haven’t tried the alarm/stopwatch features as they’re not exactly needs of mine. Overall, I like that the look of the watch I like better in comparison will not only serve its purpose for my schooling just fine, but it also looks less kiddy, was cheaper, and actually came with instructions. If all you care about is big easy to read characters and don’t mind if your watch time ends up being a little behind, this timex is fine. If you want/expect more/different than that, maybe keep looking elsewhere because you can probably find more of what you’re seeking out and possibly even cheaper than this timex.
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Reviewed in the United States on September 30, 2021