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Description
Rat NT-ProBNP 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. Cell Lysis Buffer: Gently wash adherent cells with ice-cold PBS, then trypsinize and collect cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be collected directly by centrifugation. Wash collected cells three times with ice-cold 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 freeze-thaw cycles or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and remove the supernatant for analysis. 5. Cell Culture Supernatant: Centrifuge at 1000×g for 20 minutes. Remove the supernatant for analysis or store at -20°C or -80°C, avoiding repeated freeze-thaw cycles. 6. 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. Preparation of standard gradient working solution: Add 1mL of universal diluent to the lyophilized standard, let it stand for 15 minutes to completely dissolve, then gently mix (concentration is 5000pg/mL). Then dilute to the following concentrations: 5000pg/mL, 2500pg/mL, 1250pg/mL, 625pg/mL, 312.5pg/mL, 156.25pg/mL, 78.125pg/mL, and 0pg/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each. Pipette 500uL of the 5000pg/mL standard working solution into the first EP tube and mix thoroughly to make a 2500pg/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 an N-Terminal Pro Brain Natriuretic Peptide (NT-ProBNP) 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 N-Terminal Pro Brain Natriuretic Peptide (NT-ProBNP) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat N-Terminal Pro Brain Natriuretic Peptide ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | N-terminal pro-brain natriuretic peptide (NT-ProBNP or BNPT) is a 76-amino acid N-terminal inactive protein precursor, from which brain natriuretic peptide (BNP) is cleaved. Blood levels of both BNP and NT-ProBNP are used to screen for and diagnose acute congestive heart failure (CHF) and to determine the prognosis of heart failure, as both markers are often elevated in patients with a poor prognosis. Plasma concentrations are also often elevated in patients with asymptomatic or symptomatic left ventricular dysfunction and are associated with coronary artery disease and myocardial ischemia. | |||||||||||||||||||||||||||||||||
| 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 | 78.12-5000pg/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, tissue homogenate, cell lysate, cell culture supernatant and other biological fluids |
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4.3 ★★★★★
Based on 1853 reviews
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Product Reviews
★★★★★ 5
My 90 lb. Pitbull's All-Time Fave Toy
Size: Large, Color: Coral, Size: Large, Color: Coral
I ordered a plethora of new toys for Bruce to keep him occupied while I am working remotely from home. Poor guy gets so bored while I'm stuck in front of my computer 8+ hours a day. All the other toys are animated, but this one is just a big floppy chewy squeaky toy. Bruce quickly lost interest in all the other chirping, hopping, and twirling toys and fell in love with "Diny". When I opened Diny and gave it to Bruce, he had the elated and surprised look of a 3 yr old at Christmas. And he stayed that happy for hours, and then each time he picked up Diny to play. Diny became his go-to toy over all others (and there are many!). It is well made out of durable fabric, and they did not stuff it full of kapoc (I hate kapoc in dogs toys!). And it is delightfully low-tech. Normally he tears a squeaky toy apart as soon as he can isolate the squeaky mechanism. But cleverly, the squeaky box is loose in the long belly so it moves from one end to the other as the toy is handled. That made it difficult for Bruce to locate it. The toy remained in good condition for 2 weeks of near constant attention by Bruce. But yesterday he zeroed in on the squeaky box and ripped Diny to shreds. I was sad to see Diny go so soon. But boy-howdy was Bruce happy with it. I'm gonna buy another because we liked it so much. And so did I : )
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Reviewed in the United States on May 23, 2024
★★★★★ 5
This is one “under the radar” gem!!
Size: Large, Color: Coral
This is one “under the radar” gem!! Very strong internal squeaker and way tougher than it looks. My big 125lb Shepard-Husky mix gave it heck, then made it his new buddy. He’ll occasionally give it the business but on the whole, it’s surprisingly tough and my big boy loves it. Nice price!!
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Reviewed in the United States on May 10, 2026
★★★★★ 4
Cute but small
Size: Large, Color: Skinny Green
Cute stuffy and has not yet ripped. While my dogs plays with it, the toy is a bit small for him. For reference he is 120lbs.
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Reviewed in the United States on May 27, 2026
★★★★★ 5
My Puppy Loves this Indestructible Dragon
Size: Large, Color: Pink, Size: Large, Color: Pink
My puppy (8 month old black lab mix) loves, loves, loves her dragon. She has had it for 2 months now. It is one of her favorites and gets daily rough play time. It's her go-to toy for fetch and tug games, as well as just general mouthy play. The squeaker gave out after about 4 weeks, but she would binge squeak for prolonged periods (kind of like chomping on gum) so it was almost a blessing when it went silent. She still chomps on it, but at least now I don't have to hear it. Other than the fact that it's a little grungy and bedraggled from so much loving, it is still in amazing shape. The seams and fur are all still intact. There are no visible holes, rips, or weak spots. I love Chew Guard Technology.
This dragon replaced her goDog Iguana (from PetSmart) that developed a tear in the fur after about 4 months of heavy use. (It was superficial, just the outer fur layer and not the innards, but we disposed of it before she could eat more fur.) She can be pretty rough on her toys and I can never predict if any given toy will last minutes or months. The goDog stuffies definitely fall into the "months" category, even with rough daily use.
Edit: The dragon lasted about 6 months of intensive, hard play before it developed a small tear in the fur. I patched it a couple times, but once my pup discovers a weak spot, it's the kiss of death for any stuffed item. We had to dispose of the dragon (so sad) to keep her from eating any more of the fur. We replaced the dragon with the goDog Chameleon, which is still going strong. I think we will go back to the dragon for our next toy because it's just so darned cute. We gave the dragon to my niece for her golden retriever puppy who is very chew-happy. That dragon is also doing well. You've just gotta love that chew-guard technology. This is one tough toy.
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Reviewed in the United States on September 26, 2015
★★★★★ 5
Precious toy for big dogs too
Size: Large, Color: Skinny Green
Not only is this toy cute and fun, but material is thick and durable even for my crazy GSP who is a destruction artist. This is, the favorite toy his go to in the toy box.
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Reviewed in the United States on May 15, 2026