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Description
Human TJP2 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: Gently wash adherent cells with pre-chilled PBS, then trypsinize and collect the cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be collected directly by centrifugation. Wash the collected cells three times with pre-chilled 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 freezing and thawing or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and collect the supernatant for analysis. Cell Culture Supernatant: Centrifuge at 1000×g for 20 minutes. Collect the supernatant for analysis, or store at -20°C or -80°C, but avoid repeated freezing and thawing. 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 utilizes 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 Tight Junction Protein 2 (TJP2). 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 Tight Junction Protein 2 (TJP2) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Tight Junction Protein 2 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Tight junction protein 2 (ZO-2), also known as TJP2, is a protein encoded by the TJP2 gene. Tight junction proteins (TJPs) belong to the membrane-associated guanylate kinase (MAGUK) homolog family and are involved in the organization of epithelial and endothelial intercellular junctions. TJPs bind to the cytoplasmic C-termini of transmembrane junction proteins and connect them to the actin cytoskeleton. They play a role in tight junctions and adherens junctions. They have been shown to interact with claudin 1, protein 4.1, occludin, and USP53. | |||||||||||||||||||||||||||||||||
| 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 | 0.312-20 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, cell culture supernatants, and other biological fluids |
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4.9 ★★★★★
Based on 5 reviews
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Product Reviews
★★★★★ 5
Outstanding Story!!!!!!!
Format: Kindle
Oh I loved this story. Sera is such an amazing, fun character. She has so much fire inside of herself. She has humor that makes you laugh with her sassy comments. The guys are fantastic. I don’t feel as if we really have gotten to know who and what they are but they are such a mix of fae. The story is extremely well written and developed. It had me pulled in and didn’t let go. There was so much action, drama, and suspense. Then we get to the ending and goodness there is so much going on. This ending was a major cliffhanger and I absolutely can’t wait to see what happens next.
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Reviewed in the United States on March 12, 2024
★★★★★ 5
Exceptional
Format: Kindle
I think this an exciting entertaining story different from other fantasy reverse harmen story. I love the 1st book in this series and hope it continues to weave a story of friendship, love and disappointment as well as sadness. The cliffhanger was gripping and held you in suspense that waiting until the next book was released was almost too much. I’m so glad I waited to read this series until the majority of the books were released. Katie May and Quinn Arthur’s are wonderful writers and I’m looking forward to reading more from both of them.
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Reviewed in the United States on April 23, 2025
★★★★★ 3
I don’t mind a cliffhanger,
Format: Kindle
but I dropped at least one star because of the obnoxious gloating of the author after the cliffhanger.
Seriously - I don’t understand making your readers angry because you’re smug and expecting them to keep reading your books.
I was very definitely enjoying the series. Now I have a bad taste in my mouth and mixed feelings about continuing the series.
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Reviewed in the United States on April 9, 2025
★★★★★ 4
The series as a whole so far 5/25
Format: Kindle
I read reviews before going into this book and I don't agree with one of the more harsh ones on the main trigger she had. It is stated clearly in the forward and it wasn't as blase as it was made out to be. It definitely is touched on more and hasn't just been brushed off as the series goes
I definitely would recommend reading it. It's a good series just be for-warned
I like the series as a whole. The characters are awesome I adore the fmc shes cute and adorable but also a badass. Though there are a bunch of holes for her that I feel like just got left out.
The guys are interesting and shout out to yall for not making Gage a dragon. I'm tired of the broody ones who don't wanna talk aboit what they are being Dragons. Ki is my favorite
You can definitely tell if is written by 2 different people though because the phrasing just doesn't match up and wouldn't be something people that age says. And it flip flops between them.
I feel like there's substance without substance. We are 4 books in and we don't really know much back story on literally anyone more than right under surface deep.
There are definitely favorite MMCs which is kind of disappointing since some get shoved to the wayside. Specifically both of the best friends. They're basically useless and it's made obvious as the books go on. As well as all the men are ungodly self deprecating.
I enjoy the plot line for the most part like I said I enjoy the series its different and refreshing.
I do feel like the series is being dragged out though unfortunately. And the latest cliff hanger was just meh. So hopefully the next book is the last one.
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Reviewed in the United States on May 8, 2025
★★★★★ 5
Wow!
Format: Kindle
This book was awesome! Seraphina and her family have moved to a new town. Her family is a bit... odd. She grew up learning how to protect herself from people who might hurt her. Bloodshed is a daily occurrence with her brothers and parents during their practice sessions, and it’s all fun and games unless you need to hide a body.
Sera’s family is very close, and she’s been homeschooled most of her life. But in this new town she is going to start regular school as a senior at the local high school. Unfortunately, things at her school aren’t all they seem to be. Or perhaps more than they seem to be. Sera has her own demons to deal with, and she’s terrified her new friends will learn about her weird family and other issues and drop her like a rock. It turns out they have their own secrets as well.
This story ends on a bit of a cliffhanger and I can’t wait to read the next one! This book is well written and well edited. The heroine is spunky and has a great heart and wicked sense of humor.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on November 12, 2021