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Description
Rat ZNT1 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 10 ng/mL). Then dilute to the following concentrations: 10 ng/mL, 5 ng/mL, 2.5 ng/mL, 1.25 ng/mL, 0.625 ng/mL, 0.3125 ng/mL, 0.15625 ng/mL, and 0 ng/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each. Pipette 500uL of the 10ng/mL standard working solution into the first EP tube and mix thoroughly to make a 5ng/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 capture antibody against zinc transporter 1 (ZNT1). 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 zinc transporter 1 (ZNT1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Zinc transporter 1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Solute carrier family 30, member 1 (SLC30A1), also known as zinc transporter 1 (ZNT1), is a protein encoded by the SLC30A1 gene. It regulates zinc permeation through L-type calcium channels. It downregulates not only Zn++ influx but also Ca++ influx, thereby protecting cells from excessive cation permeation. It may be involved in zinc transport out of cells. Diseases associated with it include zinc-deficiency acrodermatitis enteropathica and epidermodysplasia verruciformis type 1. | |||||||||||||||||||||||||||||||||
| 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 | 0.156-10 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, and other biological fluids |
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4.1 ★★★★★
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Product Reviews
★★★★★ 5
A beautifully written and illustrated tale of kindness, courage and (imaginary) friendship
Format: Hardcover
A beautifully written and illustrated tale of kindness, courage and friendship (even if the friend is in your imagination). This is a great picture book for building self-esteem and for helping early learners to discover ways to overcome obstacles by showing how upbeat confidence boosting self-talk is the way to go.
I adore this beautiful family and how the climb up the slide's stairs is so relatable-you just feel the endless up, up, up as you ascend, like climbing up the stairs of the high dive and looking down and just that whoosh of fear that vertakes you from the seemingly dizzying height. I love the kindness on all the adults' faces and the cute expressions from the child. The lion is magnificent. I love that it gives shape to a child’s imaginary friend and that this friend is a true safety net -which is really the child being their own safety net. The end pages are bright orange and sunny, using the cute lion face in the pattern and all the illustrations, which are predominantly blue and yellow/orange illustrations are the perfect backdrop for the tumble of dark curls on the child’s head which allow the size of the lion to appear gigantic, and yet, calm and sweet like a kitty cat. Very sweet book! I love its message of bravery, confidence building and how you can truly be your own best friend.
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Reviewed in the United States on September 3, 2023
★★★★★ 5
Sweet Story about facing your fears!
Format: Hardcover
A trusted comrade, a little girl’s lion is always there. Her lion helps her through times of stress, or if she makes a mistake, or if she is nervous. A girl and her lion, the dream team!
One day the little girl goes on a new adventure, a brand new slide arrived at the playground and she can’t wait to try it with Lion. The ladder is a bit taller than she thought and with every climb, uncertainty starts to creep in. This slide is so high! How will we ever get down? The young girl wonders. Turning back towards Lion (since this is usually the part her brave lion steps in to help) she notices that he is afraid too!
This is such a sweet story about facing your fears! I love the bright yellow and orange with blue color palette. Having a lion inside to symbolize bravery is such a wonderful way to help young readers with facing their fears. We love to teach our children that fear is a healthy response, it’s a message that our body is giving us! It is also healthy to work past that fear (although sometimes we need some help doing just that).
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Reviewed in the United States on January 19, 2024
★★★★★ 5
Love!
Format: Hardcover
A little girl feels like she can courageously tackle anything with the help of her brave lion. When she climbs a tall slide, however, and her lion becomes afraid, she must summon her own strength to overcome her fear.
I especially love the illustrations in this validating and reassuring read.
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Reviewed in the United States on October 5, 2023
★★★★★ 5
Wonderful Picture Book!
Format: Hardcover
During tonight's bedtime reading, the captivating illustrations had my toddler completely engrossed, eagerly demanding, "again, again!" The blend of fun and valuable lessons in the story is truly a standout. It brilliantly addresses concepts of fear and doubt, offering young readers a guide on how to conquer them.
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Reviewed in the United States on August 30, 2023
★★★★★ 5
Everything a Picture Book Should Be!
Format: Hardcover
Such a great picture book. Fun, vibrant illustrations. It is so nice that As Brave as a Lion is also available in Spanish. A terrific storytime, bedtime and classroom read. The book is simple but with a great message. Highly recommend.
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Reviewed in the United States on August 12, 2023