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Description
Human Cyclin D1 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 freeze-thawed. Finally, centrifuge the homogenate at 5000×g for 5-10 minutes and remove the supernatant for analysis. Cell culture supernatant: Centrifuge at 1000×g for 20 minutes. Remove the supernatant for analysis or store at -20°C or -80°C, but avoid repeated freeze-thaw cycles. Pre-Assay Preparation: 1. Remove the reagent 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 cyclin D1 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 cyclin D1 in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Cyclin D1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Cyclin D1 is a protein encoded by the CCND1 gene. The CCND1 gene encodes the cyclin D1 protein. This gene is located on the long arm of chromosome 11 (band 11q13). It is 13,388 base pairs long and translates to 295 amino acids. Cyclin D1 is expressed in all adult tissues, except cells derived from bone marrow stem cells (both lymphoid and myeloid lineages). Its overexpression has been shown to be associated with early cancer onset and tumor progression, and it can contribute to tumorigenesis by increasing anchorage-dependent growth and angiogenesis through VEGF production. | |||||||||||||||||||||||||||||||||
| 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 | Tissue homogenate, cell culture supernatant |
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4.1 ★★★★★
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Product Reviews
★★★★★ 5
Joy restored!
Format: Paperback
Now, to be honest, I've been a John & Stasi Eldredge reader for a long time. I love their books and their writing styles. But more than their skills with language is their transparency and candor. If you get the chance to meet them, you'll see that who they are in person is just how they portray themselves in their books. They're honest. They're kind--both with themselves and with others and they're consistently dedicated to telling the truth--no hype. No bait and switch.
Now, to Defiant Joy. This is a book for those who have lost joy. This is a book for those who have been blinded to the subtle creep of disappointment and cynicism that has tragically become the filter through which they see life. Let's be honest. Life can really suck sometimes. And it's both understandable and inevitable that joy is harder and harder to find...much less experience. Stasi has lived enough life to understand that condition. She shares her own struggles and disappointments--all of which give her credibility when she invites her readers to consider seeking joy in the midst of less than joyful circumstances.
"Defiant Joy" is a fitting title for those who don't want to give up yet; who don't want to give in to resignation. And that's the invitation Stasi gently offers--there is another option beside resignation, cynicism, depression and disillusionment. Joy! Who would have thought?
Stasi writes like she talks. Reading her work is like sitting down with a good friend for an honest and frank conversation about what really matters. She's the kind of friend we all long to have--one who tells us what we need to hear instead of what we want to hear; one who tells us the truth, even when we've lost hope of ever finding it. You won't find any relativism or warm-fuzzy self-talk. This is not a self-help book. Stasi has suffered too much to offer simplistic answers to some of the most difficult questions.
I highly recommend this book. It's on my suggested reading list for the community I lead. Stasi is one battle-experienced traveler who's offering other weary travelers hope for the journey ahead. I'll bet money you'll discover joy again.
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Reviewed in the United States on January 18, 2019
★★★★★ 5
You can experience joy!
Format: Paperback, Format: Paperback
What is joy? How does it compare to happiness? From where does joy come? How can we experience joy every day despite the pain and sorrow that we all experience?
Stasi Eldredge writes honestly and poignantly from places of pain and sorrow in her own life - but doesn't leave us there. She leads us into hope, into beauty and into a new depth of life in God. She teaches us how to cultivate joy in really practical - and achievable - ways.
I was especially struck by her walking through St. Patrick's Breastplate - an ancient prayer - and how I can turn from a place of failure and shame based on my performance to the King and His character and hide myself in Him.
"I need mercy, and I know it. In that knowing comes a great gift. I turn my heart again to my kind and understanding God and confess to Him that I need mercy. His answer swamps my heart with a too-good-to-be-true reality that leads to a crumbling of hopelessness and shame. My self-loathing collapses into His love. My self-condemnation melts into His arms that welcome and soothe. I have blown it. The blowing now has become the wind of the Holy Spirit. Ruah is here. His breath shepherds my heart into my Father's, and there mercy triumphs over judgment."
I encourage you to read this book. God will use it to restore your soul, to bring water to your dry and weary soul, and to help you find rest in His strong arms. You will not feel condemned, but encouraged. I found hope in its pages.
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Reviewed in the United States on October 17, 2018
★★★★★ 5
Good
Format: Paperback
Good
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Reviewed in the United States on April 15, 2026
★★★★★ 4
Excellent, inspirational, challenging
Format: Kindle
Stasi doesn't just support her vision that God wants us live joyously with scripture, she acknowledges the challenges to that, the pain we experience in life, and then gives us practical advice for applying God's love through those times to become deeper, better, more loving individuals on the other side. I wish I had read this book before my Daddy was called home, but I guess I wouldn't have been the same person if I hadn't experienced that pain alone. Defiant Joy leads you beyond surface Christianity and challenges you to see God more closely.
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Reviewed in the United States on July 18, 2019
★★★★★ 5
Love itttt
Format: Paperback
Such a great book! Everyone should read it
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Reviewed in the United States on December 5, 2025