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Description
Mouse PTGS2 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: 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: 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. Plasma: Collect specimens using EDTA or heparin as anticoagulants and centrifuge them at 1000×g for 15 minutes at 2-8℃ within 30 minutes of collection. The supernatant can be tested or stored at -20℃ or -80℃, but repeated freezing and thawing should be avoided. 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 30 ng/mL). Then dilute to the following concentrations: 30 ng/mL, 15 ng/mL, 7.5 ng/mL, 3.75 ng/mL, 1.875 ng/mL, 0.9375 ng/mL, 0.46875 ng/mL, and 0 ng/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each tube. Pipette 500uL of the 30ng/mL standard working solution into the first EP tube and mix thoroughly to make a 15ng/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 prostaglandin endoperoxide synthase 2 (PTGS2) capture antibody. 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 prostaglandin endoperoxide synthase 2 (PTGS2) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Mouse | |||||||||||||||||||||||||||||||||
| Synonym | Mouse Prostaglandin Endoperoxide Synthase 2 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Prostaglandin Glycogenase Synthase 2 (PTGS2), also known as cyclooxygenase-2 or COX-2, is an enzyme encoded by the PTGS2 gene. It converts arachidonic acid (AA) into prostaglandins (PGs). It is a sequential homodimer. Each monomer of the enzyme has a peroxidase and a PTGS (COX) active site. PTGS (COX) catalyzes the conversion of arachidonic acid to prostaglandins in two steps. | |||||||||||||||||||||||||||||||||
| 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 | 0.46-30ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, and other biological fluids |
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Product Reviews
★★★★★ 5
Great Read, and Awesome Resource.
Format: Paperback
Great Book to give insight to the blue team side.
It has some of that simple common sense stuff. But it takes that and puts it into a plan!
Very deep insight into the importance of being prepared and explaining why.
Also has some cool more advance advice to read up on.
Gives command line examples of different parameters to use and shows the result you will get.
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Reviewed in the United States on November 30, 2016
★★★★★ 5
Original review was 4 stars...
Format: Paperback
Simply because I didnt know what I was doing. This manual will grow with you. I have added and taken away some tips and tripsnof my own. Awesome read!
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Reviewed in the United States on October 17, 2017
★★★★★ 5
Great Resource to Have on hands for Security Analysts
Format: Paperback
Great Resource to Have on hands for Security Analysts. I have been using many of the techniques described in this book over the past few years but this book expanded on that knowledge a bit more than I was used to. It's great because it has all of the most common things one would typically run into on their day-to-day job as a security analyst. Not completely comprehensive but that's not the purpose of this book.
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Reviewed in the United States on September 26, 2015
★★★★★ 4
Great book
Format: Paperback
Nice companion to the RTFM. I have been in INFOSEC for a while and I found most of the book to be somewhat generic. My feelings may not apply to everyone else though. There seems to be an idea out there that everyone in INFOSEC sits around grepping Snort logs and/or TCP dumps; this simply is not the case. As an incident first responder the job is fairly straight forward: isolate and collect the logs. Everyone has their own methods I guess. Some OS'es require you leave the system connected before collecting logs this book does not cover that aspect. Not everyone can afford a SANS course so in that respect this condensed field guide is a plus+. Simply put; you cannot afford to not have this book. A must buy.
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Reviewed in the United States on October 11, 2014
★★★★★ 5
Five Stars - Version 2 update very nice
Format: Paperback
Update - version 3.0 has soooo much more depth. Really nice update.
Solid info, brief and informative, wide coverage on incident issues.
Bought a second copy recently (gave away #1), have read through it, and the update does improve wording, fixes the few spelling errors someone complained about in V1, and has great packet header charts in the back. Got my copy autographed at a SANS conference to boot! I hear there are more titles in the series planned with a few underway.
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Reviewed in the United States on August 31, 2014