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Description
Rat ND1 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 NADH-ubiquinone oxidoreductase chain 1 (ND1) capture antibodies. 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 NADH-ubiquinone oxidoreductase chain 1 (ND1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat NADH-ubiquinone oxidoreductase chain 1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | NADH dehydrogenase 1 (NADHD1), also known as ND1 or MT-ND1, is an enzyme encoded by the MT-ND1 gene. This protein is a subunit of NADH dehydrogenase, located on the inner mitochondrial membrane and the largest of the five complexes in the electron transport chain. It is a subunit of complex I of the respiratory chain and is considered to be part of the minimal assembly of core proteins required to catalyze the dehydrogenation of NADH and electron transfer to ubiquinone (coenzyme Q10). | |||||||||||||||||||||||||||||||||
| 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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4.3 ★★★★★
Based on 12 reviews
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Product Reviews
★★★★★ 5
Emotional read with children
Format: Kindle
Cammie is a single mother of two children whose ex-boyfriend, who is an alpha that he met his scent matched omega and dumped her via messenger on his phone then the next day put their house up for sale. Cammie moves back to her hometown, but because she has no support, is working two jobs to keep her and the children in a small rented house. One night her four year gets out of the house and she meets a neighbour, but she refuses to make the same mistake again by getting involved with another alpha, but Reid is convinced they are scent matched mates even though Cammie is a beta. Reid and his omega Fin start to court Cammie and her two children that they belong together as a pack and family. Cammie's ten year old son is a tough one to convince, but the scenes between him and Reid and Fin, they were so good. I was tearing up at times, found family was a big theme and I really liked this book. I highly recommend that you try it too.
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Reviewed in the United States on February 16, 2025
★★★★★ 5
It’s an experience
Format: Paperback
⭐⭐⭐⭐⭐
**A Riveting and Emotionally Charged Journey**
"Unwanted" by Sirena Song is an absolute masterpiece that grips you from the very first page and doesn't let go. The storytelling is impeccable, weaving together a tapestry of raw emotion, suspense, and heart-wrenching beauty. Song's ability to delve deep into the psyche of her characters makes them come alive, making you feel their pain, joy, and struggles as if they were your own.
The plot is brilliantly crafted, full of unexpected twists and turns that keep you on the edge of your seat. Each chapter unfolds like a piece of a complex puzzle, gradually revealing the depth of the narrative. The themes of love, rejection, and redemption are explored with such nuance and sensitivity that it leaves a lasting impact.
Sirena Song's prose is poetic and evocative, painting vivid pictures with her words and creating an immersive reading experience. The character development is outstanding, making it easy to become emotionally invested in their journeys. The ending is both satisfying and thought-provoking, leaving you pondering long after you've turned the last page.
"Unwanted" is not just a book; it's an experience that touches the soul. I highly recommend it to anyone looking for a powerful and unforgettable read. Sirena Song has truly outdone herself with this one.
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Reviewed in the United States on June 30, 2024
★★★★★ 4
Totally Sweet
Format: Kindle
🔥OmegaVerse
🔥Single Mom
🔥Scent Match
🔥Emotional Trauma
Well written, emotional omegaverse about a single mom of 2 with an ex-Alpha boyfriend that found his Omega and ditched his family. Cammie has a meet cute with Reid and Finton separately but they both find her to be their scent match.
This book is totally sweet with lots of drama but lots of proving to Cammie that she is loved and wanted.
My Personal Ratings:
5 Stars: Loved it! Couldn’t put it down or obsessed about it while needing to get to the end of it. Had to share some bits with my hubby!! Usually read in less than 2 days cause a girl has got to sleep sometime. Will be reading everything this author has to offer and most likely reread the series as next books in the series are released.
4 Stars: Really Good. Whether the story drew me in or a specific character, will definitely continue the series and look into more books by this author.
3 Stars: Enjoyed something about the book, whether it be the storyline or a character but also may have not liked something. Sometimes the book is great but needs some TLC from an editor. May or may not continue the series.
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Reviewed in the United States on July 5, 2024
★★★★★ 5
Read it!!! It's so good!
Format: Kindle
I will reccomend this book to anyone and everyone who is looking for an omegaverse book to read! The rabbit hole of male omega books this one sent my mood reading self down was crazy! 🤭 I could not have loved Finn more! He was the sweetest omega, making sure Cammie was comfortable each step of building a relationship with him and Reid. He was so good with the kids even though Ben was a little rough around the edges.
Cammie's fears were all justified but the way she learned to trust her new pack was sweet! They just continued to show up and her sister was hilarious! Being a mom and thinking of her littles but also finding her own happiness was such a sweet journey for her 💖 lets just talk about the kids for a second and say Em is a spitfire and I LOVE her and Ben sweet Ben just wants to protect his Mama but needs love, Reid does such a good job stepping up!
And the 'gift' at the end had me in tears!! 🥹 Without giving too much away that is why I think sweet Finn is my favorite of all male omegas I have ever read! His way with words no matter what he is using them for, declarations of love, pep talks, or advice he really just knows what needs to be said 💖
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Reviewed in the United States on May 31, 2025
★★★★★ 3
I wanted to like this more than I did
Format: Kindle
This started off so well! I thought it was going to take Lola on a run for her money but it ended up getting weird. I liked the writing but there was just too much spice which I didn't think was possible for me. There were too many times that the writing felt weird or the situations too outlandish and unrealistic. I had to skim through a lot of the later half. It might just be the dynamic between the characters that didn't work for me so I'll give this author another go.
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Reviewed in the United States on February 7, 2025