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Description
Human TPM4 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 tropomyosin alpha-4 chain (TPM4) capture antibody. After incubation and washing, the sample 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 tropomyosin alpha-4 chain (TPM4) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Tropomyosin alpha-4 chainELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Tropomyosin alpha 4 (TPM4) is a protein encoded by the TPM4 gene. TPM4-associated diseases include inflammatory myofibroblastoma and autosomal dominant thrombocytopenia. Pathways involved include Rho GTPase signaling and ALK signaling in cancer. Gene Ontology (GO) annotations associated with this gene include calcium binding and muscle structural components. An important homologue of this gene is TPM2. | |||||||||||||||||||||||||||||||||
| 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 ★★★★★
Based on 8 reviews
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Product Reviews
★★★★★ 5
I read the quotations first
Format: Hardcover
Of all the stories I have read about those who experienced extraterrestrial life, and all those who told those stories, Billy Meier's story is the most authentic. This book is a beautifully written and photographed narrative of Meier's relationship with various people from the stars, over a period of time. Billy himself reminded me of my best friends in the 1960's and 70's--quiet and kind. But the most remarkable part of this book, apart from the extraordinary photographs, is that there are quotations from the extraterrestrial visitors themselves, which are worth reading, in order. Yes, you might still not believe this happened, but you cannot help but absorb insights in those quotations. You will not forget them.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on July 17, 2024
★★★★★ 5
A Civilization in Transition: Can Humans Make the "Jump" to Hyperspace?
Format: Hardcover
Though there is ample evidence to suggest that humanity has been visited by "off-planet" beings or relatives since as far back as any recorded history in stone or on paper, with the advent of technologies bringing us closer to an enlightenment or a destruction, UFO/UAP events seem to becoming more frequent. And more accepted. When UFO...CONTACT FROM THE PLEIADES v I and II were first published 45 years ago few people had a personal computer in their homes let alone any digital technology whatsoever. There were gains in human rights protections along with ecological study and the awareness of the importance to "love our Mother." (Earth) So it may not be so unreal to consider that one specific human, a farmer in rural Switzerland was chosen for multiple contact events and allowed to capture photographs, audio recordings, films and even metallic samples from OP visitors for the purpose of helping humanity expand its consciousness to the next level.
This reissue tells such a story with vivid photography, detailed analysis and skilled reporting having used the most advanced non-civilian equipment available at the highest levels in its day. Furthermore, time has only affirmed that none of the photographs or video from the Billy Meier case have proved to be fake or forged in any known way.
This is the book for searchers interested in the "nuts and bolts" of ufology. And yet...
Whether you believe in UFOs or off-planet visitors is really beside the point here. The deeper point - and the not so hidden purpose revealed through the messages given to Billy Meier from Semjase, a Pleiadean traveler who contacted Meier over 100 times in the 1970s - is to consider if each one of us is capable of suspending disbelief long enough to be curious, which takes an open heart. And if we open our hearts, how can we do harm to ourselves, our planet, or others in the universe? And if we can achieve "do no harm" what else could we achieve? Hyperspace? Telepathy? A humane civilization where no one goes hungry or homeless? The point is whether or not we are ready, collectively, to expand our consciousness.
After reading this book, I want to take that leap!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on May 3, 2024
★★★★★ 5
A wonderful book !l
Format: Hardcover
No doubt the subject of UFO's and ET's is a challenging subject. But the Billy Meier story is certainly one of the very best. This book is masterwork that warrants the highest rating. The photos are extraordinary and the story line is superb. Buy a copy or several for yourself and for gifts .
A. very happy customer ....indeed.
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Reviewed in the United States on October 22, 2024
★★★★★ 5
Amazingly well written
Format: Hardcover
Nice book. Pictures and honest interviews and reports via credible down to earth persons.
They don't weigh you down with hundreds of pages to read. Well done
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on September 15, 2024
★★★★★ 5
Beyond Expectations
Format: Hardcover
Since I have had several experiences I am always looking for something legitimate -something that looks into the heart and truth of the material and comes out authentic to the core - I love this book - It far exceeds what I expected and am very happy I ordered it to add to my collection. This will be on the top of the stack! The only down side is that amazon packaged it rougly where it slid freely about in an overlarge box --This ended up tearing the top of the dj rear --- :( I do hope they do not pack all orders for this bk that way - if there are notes you can add to your order you might request book packaging or something more durable than sliding loosely about I found out about this book on the newsletter I receive from Shirley MacClaine - she always has valuable nuggets to share !!!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on April 23, 2024
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