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Description
Rat PF-4 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 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. 3. Cell culture supernatant: Centrifuge at 1000×g for 20 minutes. Remove the supernatant for testing or store at -20°C or -80°C, but avoid repeated freeze-thaw cycles. Pre-Assay 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 stand for 15 minutes to completely dissolve, then gently mix (concentration 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 seven EP tubes and add 500uL of universal diluent to each tube. Pipette 500uL of the 20ng/mL standard working solution into the first EP tube and mix thoroughly to make a 10ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube is used 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 Platelet Factor 4 (PF-4) 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 Platelet Factor 4 (PF-4) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Platelet Factor 4 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Platelet factor 4 (PF4) is a specific protein synthesized by platelet α-granules. It is a tetramer of basic polypeptides. It readily binds to and neutralizes heparin and readily binds to heparan sulfate on the surface of vascular endothelial cells, slowing thrombin inactivation and thereby promoting thrombosis. It is a small cytokine belonging to the CXC chemokine family and is also known as chemokine ligand 4. | |||||||||||||||||||||||||||||||||
| 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 | Serum, plasma, and cell culture supernatant |
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4.3 ★★★★★
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Product Reviews
★★★★★ 5
Patience and Christianity
Format: Kindle
Short review: buy it
Long review:
It is incredible the simplicity of the Christians praxis in its origins, and how Saint Augustine and then Luther totally misunderstood these origins.
I can’t give you all the thoughts about this book, but here a glimpse of some ideas:
- The forgiveness between Christians is still powerful mean to live in peace and in a productive way
- The peace kiss is now forgotten, but it was a very powerful practice that maintain unity in the communities
- Women in the church were extremely important, they helped with maintain the union and share information
- The first Christian didn’t think that mission was most important than behavior, and for good reasons: talk is cheap, actions are more important.
- The testimony was noting about believe, it was about behave as a Christian, you can only access the great teachings of the New Testament once you showed with your actions that you are worthy of that.
Incredible simple, I think that is difficult to destroy religion only with reason, because religions have nothing to do with theology, is about behavior and cooperation.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on November 17, 2020
★★★★★ 5
Almost Persuaded
Format: Kindle
Almost thou hast persuaded me to become a pacifist. Kreider presents an outstanding survey of the Christian emphasis on patience in the first four centuries of the church. I am rethinking everything. Loved this book.
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Reviewed in the United States on November 27, 2023
★★★★★ 5
A great handbook for letting go of toxic relationships!
Format: Paperback
I wish I had read this ten or twenty years ago, but its never too late! Even as a grandmother I am always trying to improve relationships, and only noticed recently that I needed to leave many old acquaintances out of my contacts when I got a new phone number. If they always drained me when we talked or if I had a hard time getting to a positive place after a conversation with them, I just let them go. I loved seeing the ways Jesus walked away too! Very good information at any age and permission granted to spend your time on the people that fill you up, not take you down their dark hole where everything is negative and hopeless. Also helps continue to deal with people you can't walk away from and save your sanity!
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Reviewed in the United States on March 28, 2025
★★★★★ 5
Excellent Information on how to stay focused on our spiritual path!
Format: Paperback
I accidentally purchased this book. I intended to get the actual book instead of the study guide.
I did order the actual book and it is incredibly helpful!
As a person in recovery for codependency, I have struggled all of my life with being nice and respectful to everyone no matter how I was treated. It is exhausting, and created so much anxiety and uncertainty. The insight from this book, has shown me how to stay calm , deliberate and focused on my spiritual path.
In recovery, Step 3, we are invited to make a decision to turn our will and life over to the care of God. This book has shown me what has been missing in my recovery. I have allowed toxic people to distract, belittle, humiliate me all in the name of being a Christian, and tolerating everyone and their toxic behavior. Even people in the church! This information has validated my need to be mindful and be strong in my faith, as I live out Gods plan for my life. I have put people before God...yep. No one is more important to me than God. Now I can live my truth and be more effective in Gods will for my life.
It truly seems to be my healing journey of putting God first in my life. It is a decision that is at the core of my existence! Thank you Gary Thomas for addressing the concerns of evil on our path and to be resilient and follow only that still small voice and silence the loud, obnoxious voices of those who add nothing to our life. Life giving read for me!
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Reviewed in the United States on December 26, 2020
★★★★★ 5
Great Book, Life Lessons
Format: Paperback
We did this study with our small group. We all have similar backgrounds and this study was a great way for us to work through some of difficult relationships . We learned how to support one another and how to draw healthy, biblical boundaries within the difficult relationships in our lives.
I’ve done other Gary Thomas studies before, and once again he writing does not disappoint . His teaching is easy to follow, steeped in scripture, and full of life application.
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Reviewed in the United States on April 13, 2024