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Description
Human STAT4 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 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 Signal Transducer and Activator of Transcription 4 (STAT4) capture antibody. After incubation and washing, the assay 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 Signal Transducer and Activator of Transcription 4 (STAT4) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Signal Transducer And Activator Of Transcription 4 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Signal transducer and activator of transcription 4 (STAT4) is a transcription factor belonging to the STAT protein family, which consists of STAT1, STAT2, STAT3, STAT5A, STAT5B, and STAT6. STAT proteins are key activators of gene transcription, binding to DNA in response to cytokine gradients. STAT proteins are common components of the Janus kinase (JAK) signaling pathway and are activated by cytokines. STAT4 is required for the development of naive CD4+ T cells into Th1 cells and for the production of interferon-γ in response to IL-12. There are two known STAT4 transcripts, STAT4α and STAT4β, which differ in their downstream interferon-γ (IFN-γ) production. | |||||||||||||||||||||||||||||||||
| 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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4.3 ★★★★★
Based on 27 reviews
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Product Reviews
★★★★★ 5
Durable toy for a great price! Will buy again.
Color: A Red, Size: For Larege Dogs, Color: A Red, Size: For Larege Dogs
Great toy, heavy, good size, and durable for active chewers. My two dogs like to play tug a war a lot and this toy has held up fantastically! Afterwards they tend to chew toys to pieces but this one has stood the test of time so far. Great buy!
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Reviewed in the United States on May 14, 2026
★★★★★ 4
Very tough & VERY heavy
Color: A Red, Size: For Larege Dogs
My dog is a half black lab/half pittie & a very aggressive chewer she goes through toy bones very quickly, to the point they're so slivered up within 2 days, sometimes slightly longer, I have to get rid of them, they never last long, except for whole antlers and some split antlers. Although she rarely touches the whole antlers . Anyways, this is a great tough toy for strong large breeds. It squeaks in the center if you squeeze hard enough but the toy/bone was a bit too heavy for my girl. I got her the next smaller one and she still plays with the big heavier one more than the one that fits her perfectly so go figure. It's a great product and definitely recommend for heavy chewers. I only took a star off because of the weight I was extremely surprised by how heavy it was, but that's not necessarily a bad thing depending on the dog, this size is just not for my dog. I think I've had it about a month and she does play with it, not as much as her other toys, but it's still in amazing shape even with her heavy chewing.
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Reviewed in the United States on April 21, 2024
★★★★★ 5
Squeaky Toy for Aggressive Chewers
Color: A Violet, Size: For Larege Dogs
Size is great for our big dog, he loves to chew & squeak it. engagement, our dog loves playing fetch, hours of enjoyment.
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Reviewed in the United States on May 21, 2026
★★★★★ 3
Maybe not for super aggressive chewers
Color: A Violet, Size: For Larege Dogs, Color: A Violet, Size: For Larege Dogs
I had high hopes for this toy, I was so excited about it. I have aggressive chewers, two Malinois/Dutch Shepard mixes and a Pit. The Shepard's chewed pieces off of it in literally 5 minutes. At the ends there is a small lip and that's where they started chewing pieces off. I had ordered a second one before I gave them this one (early Christmas present) because when I received it I noticed it felt very sturdy and durable. They do like it so since they already chewed this one I'm going to see if I can shave that lip off and maybe they won't get anymore pieces off. That'll decide if I return the one that arrives tomorrow. Maybe not good for super aggressive chewers. It's very thick and seems well made. Might just be better for other dogs.
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Reviewed in the United States on December 21, 2025
★★★★★ 5
Fantastic for rough chewing
Color: Bule 0, Size: For Larege Dogs
Our big boy is rough with his toys and always seems to disintegrate his toys in seconds. I had seen this with hoping this will be the toy he doesn’t destroy. On day 2 now with the toy and he’s obsessed, even feel asleep with the toy in his mouth and still in one piece. Highly recommended for tough chewers.
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Reviewed in the United States on May 25, 2026