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Description
Human PAR4 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 capture antibody against Protease Activated Receptor 4 (PAR4). 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 Protease Activated Receptor 4 (PAR4) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Protease Activated Receptor 4 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Protease-activated receptor 4 (PAR4) is the fourth member of the PAR family and the third thrombin receptor identified and cloned in 1998. PAR4 is expressed in platelets and several other human tissues, such as the lung, pancreas, thyroid, testes, and small intestine, but is absent from the brain, kidney, spinal cord, and peripheral blood leukocytes. PAR4 comprises 385 amino acids and shares 27% and 30% sequence identity with PAR1 and PAR3, respectively. | |||||||||||||||||||||||||||||||||
| 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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4.2 ★★★★★
Based on 9 reviews
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Product Reviews
★★★★★ 5
They really do work!
Color: Zero Pore Pads, Size: 70 Count (Pack of 1)
I rarely give reviews that aren’t motivated by disappointment, but this is one of them. Medicube makes several products that I use alongside $170 serums and creams, and the Zero Pore Pad is one of them. I love the dual texture, and even after using a pre-cleanser and a “microfolient” face wash (both expensive washes from by Dermologica), the Zero Pore Pad is my ONLY choice for a toner. Compared to the products that go on after using the pads, the pads are incredibly affordable for the results I’ve seen. On the rougher side of the pad, this toner still somehow finds pockets of sebum that are visible on the pad after using as directed. It’s like magic.
I’m on my third container of Zero Pore Pads and I can say, without a doubt, they have had noticeable results on my skin, mostly on my nose and the cheek area around my nose. The size of my pores are now smaller, almost invisible. This is despite the fact that I layer a bunch of serums, creams and a moisturizer at night that I lock in with the peel-off Medicube collagen mask. Despite the weight of it all on my pores, the pads still have worked, and they aren’t harsh like some toners-but if you have very sensitive skin or are on acne medication, go with their milder sister product). I wash my face three times a day due to the ick of polluted air and layers of moisturizer and sunscreen, so I’ve been using these three times a day, but occasionally I use the Koji and Tumeric Pads, which have had no noticeable effect aside from serving as a basic toner.
The application is lovely on all Medicube’s pads with their dual texture, but like the collagen mask these Zero Pore Pads really do work (for me). Because skin varies with age and melanin content, I am a pale white woman (female at birth) of Scandinavian descent so I don’t know if these would have had the same impact on me at age 17, but now they work, really well. I literally have the glass skin K-beauty prizes—I just used the pads and thought I had something on my face because my skin was shining. I love these, try them!
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Reviewed in the United States on March 29, 2026
★★★★★ 4
Good toner pad option
Color: Zero Pore Pads, Size: 70 Count (Pack of 1)
I like these to use no more than 1-2x a week for my skin. I use them before bed and wake up noticing smoother and more event toned skin. If you use them too often, they can cause redness and irritation, but I have never gotten a rash or a breakout because of them. I will say, they do not exfoliate all your texture away and refine your pores to perfection like the marketing says, but nothing does. The AHA and BHA are the chemical exfoliants and work just like they do in any other product. The pads are very moist and stay saturated in the container as long as you keep the lid shut. Overall, a good toner pad option to be used occasionally in your routine and do not mix with any other actives.
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Reviewed in the United States on May 12, 2026
★★★★★ 5
Ive been ordering these for years now
Color: Zero Pore Pads, Size: 70 Count (Pack of 1)
I’ve been using these toner pads twice a day and they’ve worked really well for my skin. Since using them consistently, my acne has stayed under control much better than before. The pads feel gentle and don’t irritate my skin at all. I also like how convenient they are compared to using separate toner and cotton pads. My skin feels cleaner and smoother after each use. The container stays sealed well so the pads don’t dry out quickly. Overall, these have become a staple in my skincare routine.
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Reviewed in the United States on May 21, 2026
★★★★★ 5
The Biggest Visible Change I’ve Seen in My Skin in Years
Color: Zero Pore Pads, Size: 70 Count (Pack of 1), Color: Zero Pore Pads, Size: 70 Count (Pack of 1)
If I could only keep one skincare product, it would honestly be these pads.
Even after trying peels, tools, clay masks, exfoliants, and retinol, this is the most noticeable improvement I’ve seen in my skin.
I’m in my 50s and have tried a lot of skincare over the years — pore strips, clay masks, exfoliants, retinol, you name it. Very few products have made a noticeable difference in my skin, but these absolutely have.
My biggest concern has always been the pores on my nose. They’ve looked enlarged for years and it’s something I’ve been self-conscious about. After using these pads, the difference was surprisingly visible — my pores looked tighter, my skin texture smoother, and my nose looked noticeably clearer.
I added a close-up photo because this is where I’ve seen the biggest difference — my pores look visibly tighter and clearer
What impressed me most is that I saw improvement right away, but the results actually kept getting better with consistent use. After about a month, my skin looks more refined overall and my nose pores appear much smaller than they used to.
I also have sensitive, combination skin, so I’m careful with exfoliating products. These have been very gentle for me. The pads are thin but well saturated (not dripping), and I like using the textured side first to exfoliate, then flipping to the smooth side to finish. There’s no strong fragrance and they haven’t irritated my skin.
Another unexpected benefit is how much better my makeup applies now. Foundation sits more smoothly and doesn’t settle into my pores the way it used to.
After years of trying different skincare products, this may honestly be the one that has made the most noticeable change in my skin.
If enlarged pores on your nose bother you like they did me, this product is absolutely worth trying.
This is one of the few products that has genuinely shocked me with how well it works! I’ve been telling everyone I know and I’m not a spokesperson! I’m just impressed and happy to find something that works so well! 🩵
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Reviewed in the United States on March 15, 2026
★★★★★ 5
Truly zero pores!
Color: Zero Pore Pads, Size: 70 Count (Pack of 1)
These toner pads are a really convenient way to incorporate gentle exfoliation into a daily routine without adding extra steps. The dual-textured design is great—the textured side helps lightly exfoliate, while the smoother side is nice for refining and prepping the skin. They feel effective without being overly harsh. With consistent use, my skin looks a bit smoother and more refined, especially in areas where pores tend to be more visible. The formula feels active, but still balanced enough for regular use if your skin tolerates exfoliating acids well.
I also appreciate how easy they are to use—pre-soaked and mess-free, which makes them ideal for both morning or evening routines. Overall, a well-formulated, easy-to-use option for maintaining smoother, clearer-looking skin with minimal effort.
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Reviewed in the United States on April 15, 2026
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