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Description
Rat TNF-b 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. Prepare the 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 300pg/mL). Then dilute to the following concentrations: 300pg/mL, 150pg/mL, 75pg/mL, 37.5pg/mL, 18.75pg/mL, 9.375pg/mL, 4.6875pg/mL, and 0pg/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each. Pipette 500uL of the 300pg/mL standard working solution into the first EP tube and mix thoroughly to make a 150pg/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 Tumor Necrosis Factor Beta (TNF-b) 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 Tumor Necrosis Factor Beta (TNF-b) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Tumor Necrosis Factor Beta ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Tumor necrosis factor-β (TNF-β), also known as lymphotoxin-α (LT-α), is a cytokine-like protein encoded by the LTA gene. It is expressed in hematopoietic cells, exhibiting antiproliferative activity and leading to cytotoxicity in tumor cell lines. As a cytotoxic protein, it plays a variety of important roles in immune regulation, depending on the form in which it is secreted. Unlike other members of the TNF superfamily, it is found only as a soluble homotrimer and, when expressed on the cell surface, only as a heterotrimer with LT-β. It has a significant impact on the maintenance of the immune system, including the development of secondary lymphoid organs. LT-α deficiency leads to disrupted gastrointestinal development, prevented Peyer's patch development, and splenic disorders. It is involved in the regulation of cell survival, proliferation, differentiation, and apoptosis. Its effects vary depending on the organ type, cancer cell type, cellular environment, sex, and duration of action during the immune response. Produced by Th1 cells, it induces changes in adhesion molecules on the surface of vascular endothelial cells and promotes their binding to phagocytes. | |||||||||||||||||||||||||||||||||
| 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 | 4.68-300 pg/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, tissue homogenate, cell lysate, cell culture supernatant and other biological fluids |
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4.6 ★★★★★
Based on 9 reviews
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Product Reviews
★★★★★ 4
It is a great wedge pillow, but takes time to get used to sleeping on your back
Color: Dark Grey & White, Size: Adjustable 9&12 Inch + 1 Head Pillow
I bought this for post-surgery, I had to sleep on my back. Nearly impossible as a lifelong side sleeper. It is memory foam, and it does seem to run hot. But after a couple of weeks, I'm adjusting to it. Back sleeping is best for saving your face from AM wrinkles and mascara smears all over pillows. (Yes I know, I should take off mascara before bed, but....). Now that I've adjusted to sleeping with it, it stays on my bed and I wake up without the creases on my face and neck! So if you are buying pre-surgery, buy in advance to adjust to a new style of sleeping! I'm keeping it!
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Reviewed in the United States on November 7, 2025
★★★★★ 5
Its worth the purchase!
Color: Black, Size: Adjustable 9&12 Inch + 1 Head Pillow
Bought this pillow because Im dealing with both GERD and neck pain so I decided to order this and hope for the best thats being said Im so glad I did because its helped me a lot with both issues.
Some extra comments: Its a little firm but not in a bad way and the head rest is comfy but I do wish I could bring it down lower.
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Reviewed in the United States on June 3, 2026
★★★★★ 3
Misleading: Pillow Itself Has Only TWO Configurations!! >:(
Color: Black, Size: Adjustable 9&12 Inch + 1 Head Pillow
I was sold on the 10-Option bit until I realized there are only 2 choices as far as flooping and flopping the pillow itself. It is 2 triangles, a large one and a small one. The large triangle IS THE SAME ANGLE ON EACH END. The small triangle IS ATTACHED TO THE CENTER OF THE LARGE TRIANGLE. That means no matter how you move the pillow itself, you're getting the EXACT SAME THING. Where YOU come in is where all these infinity "versatility" options are - whether you put it upright or whether you put it flat. That's it. All this I can read in bed! I can play on my laptop! I can watch TV! Yeah that's upright. Option 1. And all this I can put it under my legs! I can use it as a normal pillow as well! That's flat. Option 2. There is absolutely nothing special or different about this thing except the stupid pocket. Had they not made the triangle the same angles on the end, had they made a steep angle and a low angle and that center pillow detachable but still velcro - THEN there would have been more elevation options. But this, I was highly disappointed. It's like saying, My bed has 360 options! (because I can lay 360 different degrees around the bed) AND 360 MORE BECAUSE GUESS WHAT! I CAN LAY ON MY STOMACH 360 DIFFERENT DEGREES around the bed! Oh! And on my side 360 degree different ways! Wait I have a left and a right side - the OTHER side I can lay on it around the bed 360 different degrees as well! And sitting cross-legged on the bed 360 different degree directions as well! So. No. It's a firm memory foam pillow. You lay upright or lay down or you put it under your legs or on your lap. It changes 2 ways and you make it work 4 ways. That's it. Yes it smells but not too bad, not foul, just persistently. Like a light chemical smell but it didn't aggravate me (I get scent, etc. migraines). It self-inflated pretty fast. But as you can tell, I was not impressed with it in reality. :(
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on May 3, 2026
★★★★★ 5
Helps my coughing
4.8 stars. This helps keep me sleeping at a good angle to prevent night coughing. Also, it's great for relaxing and watching tv in bed.
I find that it is Goldilocks level firm - just enough to be supportive but soft enough for comfort. I wish I'd gotten the roll pillow now :-(
It's hard to make my bed look "made" with this on it - no matter what color I get. I keep it on the side & out if sight til I need it.
I do like the material & thankfully it's removable for cleaning, but I can tell it's going to wear with a lot, so I do wish it were easier to find more covers for it so I could have one on while washing one.
Overall, I like this a lot. The only thing negative is that I can see that it will "pill" & fuzz up in places with more use. I do use this every night.
One warning: As with any products like this that are suctioned close for packaging - be VERY careful. I always hold one corner of the plastic to open a little vent before I cut more to remove the pillow and let it "fluff". It would be easy to snip some of the fabric...
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Reviewed in the United States on October 3, 2025
★★★★★ 5
Helped my husband get a good night's rest after surgery.
Color: Dark Grey & White, Size: Adjustable 9&12 Inch + 1 Head Pillow
We bought this after my husband's heart surgery, and the doctors wanted him to sleep in an elevated position for a few weeks. Before this, he was just stacking some pillows behind his neck, which did not get him a comfortable night's rest. His first night using this was a game changer for him. It was the first time he was able to sleep through the night. It was just the right height, and just the right level of soft/firm. Once he's done with it, I think I'll steal it from him and use it for reading in bed. The little neck rest was super helpful too. Thanks to whoever invented this, and sells it at such a reasonable price.
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Reviewed in the United States on March 3, 2026
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