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Description
Mouse GS ELISA KitProduct Specification Usage Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High precision pipette and gun tips: 0. 5 10uL, 5 50uL, 20 200uL, 200 1000uL 3. 37 constant temperature box 4. Distilled water or deionized water Sample processing and requirements: 1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4 overnight, then centrifuge at 1000g for 20
Product Specification
| Usage |
Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High-precision pipette and gun tips: 0.5-10uL, 5-50uL, 20-200uL, 200-1000uL 3. 37℃ constant temperature box 4. Distilled water or deionized water Sample processing and requirements: 1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4℃ overnight, then centrifuge at 1000×g for 20 minutes, and take the supernatant, or store the supernatant at -20℃ or -80℃, but avoid repeated freezing and thawing. 2. Plasma: Collect the specimen using EDTA or heparin as an anticoagulant. Centrifuge the specimen at 1000 × g for 15 minutes at 2-8°C within 30 minutes of collection. The supernatant can be assayed or stored at -20°C or -80°C, but avoid repeated freezing and thawing. 3. 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 the tissue and mince it. Add the minced tissue to the appropriate volume of PBS (generally a 1:9 weight-to-volume ratio, e.g., 1 g of tissue sample to 9 mL 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 the supernatant can be assayed. 4. Cell Lysis Buffer: Gently wash adherent cells with pre-chilled PBS, then trypsinize and collect 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 them 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, the PBS volume can be reduced 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 remove the supernatant for analysis. 5. Cerebrospinal Fluid Cell Culture Supernatant: Centrifuge at 1000×g for 20 minutes. Remove the supernatant for analysis or store at -20°C or -80°C, avoiding repeated freeze-thaw cycles. 6. 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 glutamine synthetase (GS) 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 glutamine synthetase (GS) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Mouse | |||||||||||||||||||||||||||||||||
| Synonym | Mouse Glutamine synthetase ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Glutamine synthetase (GS) is an enzyme that plays a vital role in nitrogen metabolism by catalyzing the condensation of glutamate and ammonia to form glutamine. The glutamine synthetase reaction is glutamate + ATP + NH3 → glutamine + ADP + phosphate. Glutamine synthetase utilizes ammonia generated by nitrate reduction, amino acid degradation, and photorespiration. The amide group of glutamate serves as the nitrogen source for the synthesis of metabolites in the glutamine pathway. Other reactions may occur via GS. | |||||||||||||||||||||||||||||||||
| 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 | Serum, plasma, tissue homogenate, cell lysate, cerebrospinal fluid, cell culture supernatant and other biological fluids |
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4.5 ★★★★★
Based on 27 reviews
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Product Reviews
★★★★★ 4
Knot their Omega
Format: Kindle
It took me a day to really get into the book, there is a lot happening in the beginning of the story. I wish there was more of a time frame because I felt like something’s happened quickly but they were farther away than I thought so that was a little confusing to me. I felt like there was so much happening but also not enough.
As for the story itself I enjoyed it, it was different than other OV I’ve read. Bring the tissues, you will need them. They guys were great, I wish we saw more of Kai and Kenjis relationship. As for Nate…… IYKYK.
I enjoyed this book, my first from this author and look forward to reading about Kamaris story.
Book: ⭐️⭐️⭐️⭐️/5
Spice: 🌶️.5/5
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on October 24, 2024
★★★★★ 2
Has Potential, Poorly Executed
Format: Kindle
I really really wanted to enjoy this book and was looking forward to reading it. However there were just one too many convoluted plot points, sentences just did not flow nicely at times, and I was left with a lot of confusion on my end. I think it has the bones of being a great story but seems like it was rushed and I had to push myself to finish the last of the book and was left with an unsatisfying ending. It’s definitely different from other omegaverse novels I’ve read but not for good reasons. I really think it could be great but needs some serious tweaking and editing before I’d read it again.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on February 3, 2025
★★★★★ 5
Secrets, Seduction, and Rockstars!
Format: Kindle
Rockstars, secrets, and off-the-charts chemistry? Sign me up!
Cinder Blaze takes us on a rollercoaster of passion and peril with Knot Their Omega. Blair Vesper, a secret Omega masquerading as an Alpha, strikes a risky deal to tour with Blooming Salvation, a band teetering on the edge of chaos. Enter Icarus Morrigan, the enigmatic manager, and his three complicated and irresistibly sexy rockstars: wild Kenji, icy Kaiser, and fiery Nathaniel.
This book delivers steamy Omegaverse drama, sizzling slow-burn romance, and just the right dash of angst. The tension between Blair and the band crackles like electricity onstage, while the societal stakes add depth to the spicy dynamics.
Short, sharp, and oh-so-sinful Knot Their Omega will have you hooked from the first note!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on December 9, 2024
★★★★★ 5
A good read
Format: Kindle
A good read, just fluffy cuteness, no antagonism. I like all the characters. It could have used another round of editing however, glanfds being one error that cracked me up, and my personal pet peeve was that the author kept using the word fill instead of feel, which I promise you are not interchangeable haha, but it's definitely better than the majority of books I read on here mistake-wise.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on November 28, 2024
★★★★★ 4
amazing
Format: Kindle
Knot the Bride was a fantastic read! The characters were all amazing and well-developed. It was easy to like them all. Sophia, Luca, Nick, and Gavin were all perfect for each other. It was such a charming story that had me hooked the entire time. I did wish there were POVs from Luca, Nick, and Gavin but it was still an amazing book without it.
I am excited to read the next book in the Willowside Omegaverse series! This is definitely a must-read for fans of omegaverse romance!.
I received a free copy of this book and am voluntarily leaving a review.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on April 25, 2025
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