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Description
Mouse DEFa6 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: Serum, plasma, 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
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: Serum, plasma, 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 freeze-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 freeze-thaw cycles 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 1000×g for 20 minutes, and collect the supernatant for analysis. Preparation for the Assay: 1. Remove the reagent kit from the refrigerator 10 minutes in advance and equilibrate to room temperature. 2. Prepare a gradient standard working solution: Add 1 mL of universal diluent to the lyophilized standard. Let stand for 15 minutes to completely dissolve, then gently mix (concentration 1000 pg/mL). Then dilute to the following concentrations: 1000 pg/mL, 500 pg/mL, 250 pg/mL, 125 pg/mL, 62.5 pg/mL, 31.25 pg/mL, 15.625 pg/mL, and 0 pg/mL. Serial dilution method: Take seven EP tubes and add 500 μL of universal diluent to each tube. Pipette 500 μL of the 1000 pg/mL standard working solution into the first EP tube and mix thoroughly to make a 500 pg/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 Defensin Alpha 6/Paneth Cell Specific (DEFa6) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by HRP peroxidase and to yellow by acid. The intensity of the color is positively correlated with the amount of Defensin Alpha 6/Paneth Cell Specific (DEFa6) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Mouse | |||||||||||||||||||||||||||||||||
| Synonym | Mouse Defensin Alpha 6; Paneth Cell Specific ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Defensin α6, Paneth cell (DEFA6) is a protein encoded by the DEFA6 gene. DEFA6 is expressed in Paneth cells of the ileum. α-defensins are a family of microbicidal and cytotoxic peptides that protect the host against bacteria and viruses. However, DEFA6 provides protection against invading enteric bacterial pathogens by self-assembling into cellulose and nanomembranes that surround and entangle bacteria. Several α-defensin genes, including DEFA6, are clustered on chromosome 8. | |||||||||||||||||||||||||||||||||
| 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 | 15.6-1000 pg/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, tissue homogenate, cell lysate and other biological fluids |
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★★★★★ 5
Cute jeans
Size: 12, Color: Washed Blue, Size: 12, Color: Washed Blue
Bought these jeans for my daughter and they fit great! The material is thick, which is nice even in warm weather. They look just like the photo and have a great length to them. Very comfortable and stretchy . She loves them!
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Reviewed in the United States on October 16, 2025
★★★★★ 5
FANTASTIC FIT
Size: 16 Plus, Color: Washed Blue
I'm a 4'7" , 65 year old woman. Due to abdominal issues I needed jeans that give that extra something on the waist. I've tried on just about every brand of jeans ranging in size from adult/junior 6 to 12. If they fit on the legs the waist was huge. If they fit on the waist the legs and butt had enough room for me and a friend. I was resigned to wearing yoga pants and leggings with big shirts. My son mentioned that he loved the Amazon basics men's dress slacks, so I figured why not. First I looked at women's and juniors, and I knew from the measurements that I was still stuck. Then I remembered that about 15 years ago I'd buy girls pants (I was a size 12 in kids at the time) Looking at the sizes I knew a 12 was way too small, then I saw the 16 plus. With hesitation I bought them. Well glory be miss Agnes, THEY FIT. Comfortable, real jeans again. Sure, I have to take them up about 6 inches, but that's fine, not a problem. I'm just so happy to have real jeans again !!!
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Reviewed in the United States on November 30, 2023
★★★★★ 5
Love them!
Size: 7 Slim, Color: Washed Blue
I did but expect these jeans to fit so well and look so nice on my daughter. Hands down the best jeans I’ve purchased for her, much better fit and quality than the other stores! The length is perfect, but it’s a little snug in the waist. Still a great fit with some stretchiness! Great value!
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Reviewed in the United States on July 25, 2025
★★★★★ 5
Great pair of Jean and great price!
Size: 14, Color: Washed Blue, Size: 14, Color: Washed Blue
I can’t believe these were only $13. They feel and look like quality jeans, very basic/simple boot cut. My daughter says they’re stretchy and comfortable. I ordered her a size 14 (12’s fit her waist but are always too short) She’s 11 years old, 5’ tall, about 80lbs. The waist adjusters were great for the perfect fit.
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Reviewed in the United States on August 9, 2025
★★★★★ 5
Love the Options
Size: 10 Slim, Color: Medium Wash
I love that there are slim and plus sizes. Allows me to find the perfect fit for my girls!!
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Reviewed in the United States on May 9, 2026