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Description
Human STAT1 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 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 Signal Transducer and Activator of Transcription 1 (STAT1) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by HRP peroxidase and then to yellow by acid. The intensity of the color is positively correlated with the amount of Signal Transducer and Activator of Transcription 1 (STAT1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Signal Transducer And Activator Of Transcription 1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Signal transducer and activator of transcription 1 (STAT1) is a transcription factor encoded by the STAT1 gene. It is a member of the STAT protein family. All STAT molecules are phosphorylated by receptor-associated kinases, leading to activation. They then dimerize into homo- or heterodimers and translocate to the nucleus to function as transcription factors. Specifically, STAT1 can be activated by several ligands, such as interferon-alpha (IFNα), interferon-gamma (IFNγ), epidermal growth factor (EGF), platelet-derived growth factor (PDGF), interleukin-6 (IL-6), or IL-27. Type I interferons (IFN-α, IFN-β) bind to their receptors, initiating signaling through kinases that phosphorylate and activate the Jak kinases TYK2 and JAK1, as well as STAT1 and STAT2. STAT molecules dimerize and bind to ISGF3G/IRF-9, a complex of interferon-stimulated gene factor 3 and interferon regulatory factor 9. This allows STAT1 to enter the nucleus. It plays a key role in the expression of many genes that contribute to cell survival, viability, or pathogen response. | |||||||||||||||||||||||||||||||||
| 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 | Tissue homogenates, cell lysates, and other biological fluids |
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4.8 ★★★★★
Based on 29 reviews
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Product Reviews
★★★★★ 5
An action-packed dark romantasy
Format: Kindle
I loved this book! Queen of Roses is an Arthurian-inspired dark romantasy that is the first book in the Blood of Fae series. The story follows Morgan, the princess of Camelot who is rumored to be part fae. Fueled by prejudiced hatred and a mistrust of fae blood, Morgan’s abusive father strips her of her birthright and hands it to her half-brother, Arthur. Instead of becoming queen, Morgan is commanded to join the temple of the goddesses when she comes of age.
However, Arthur turns into a psychopathic, power-hungry, fae-hating king as he ages. He develops malevolent plans and commands Morgan to find an ancient weapon with legendary power. Although Morgan is wary of Arthur’s intentions, she embraces the opportunity to go on a journey and potentially change her fate.
The story picks up from there and we follow Morgan on her quest to find the ancient relic. It’s full of high stakes adventure, mystery, tension, banter, forced proximity, hidden magic, self discovery, and betrayal. This first installment of the series intricately develops the world building and character development. There’s little romance in this book, but it is evident that it is a slow burn that will continue to develop throughout the remainder of the series.
Overall, I loved the world building, the epic fantasy, Morgan’s journey of self discovery, and all of the twists and turns that set the stage for the future installments. I can’t wait to see what happens next!
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Reviewed in the United States on April 7, 2024
★★★★★ 4
Arthurian Fae Quest…say less.
Format: Kindle
A fae centered Arthurian tale unlike any I’ve read so far. The author did a great job at descriptive world building, with scenes easily playing out in my minds eye. There was plenty of action, suspense, and even a touch of horror. An enemies to lovers, slow burn romance, a quest, with plot twist and turns aplenty. There was a love triangle, which I’m not usually a fan of but, it played out well in this story line. The FMC, Morgan Pendragon, was so blatantly naïve, yet I typically expect as much in a ‘book one’ of a series, especially one that features a fairly sheltered princess. I was happy to read that in spite of this, she still showed a strong sense of morals, fire, and spine. Now our MMC? Kairos Draven, aka Void’s Edge. Oh, how I’m a sucker for a smoking’ hot grumpy warrior alpha with a witty mouth, and a strong sense of “touch her and die” attitude, so you know who held all my cards. That ending? Just made me swoon all the harder. Now add a battlecat that rivals the size of a horse…and well Ms. Briar Boleyn you have well and truly stolen my heart. I’m excited to see where the story goes from here, and follow along to see more of the characters growth. I went into this story fairly blind, and I think I enjoyed it all the more because of it.
Once the story got going, it had me in an absolute chokehold and it was difficult to put down.
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Reviewed in the United States on May 12, 2024
★★★★★ 3
Not a bad start
Format: Kindle
3 stars
Thank you Netgalley and Briar Boleyn for the ARC!
A camelot/king Arthur retelling with fae. I was hooked by the idea of this book immediately and was eager to jump into this world.
• slow burn
• enemies to lovers
• who did this to you
Morgan Pendragon watched her mother die by her father's hand when she was just eight years old, hiding under the bed. Morgan is believed to have the tainted blood of the fae in her veins and is cast aside so that her fathers illegitimate son, Arthur, can become the king. She's seen his cruel treatment of the fae firsthand, so when he sends her on a journey to find a fae weapon she seizes the opportunity to do more with her life. Along the way, she finds more than she could have imagined.
I don't know a whole lot about King Arthur and Camelot but I had a lot of fun with this story! The plot has some similar tropes to popular romantasy books (From blood and ash) but there's enough originality here that it doesn't feel like I'm reading a copy. I liked how the fae were different in appearance than what is typical in most fantasy books I've read. In this book they have blue hair, violet skin and a wide range of other characteristics. I thought that the world building was easy to follow and I could easily immerse myself into this world. After reading the blurb I kept wondering when she was going to go on the journey to find Excalibur and it doesn't happen until around the 45% mark.
The story is a bit slow at times but starts to pick up once they begin their journey to find Excalibur. The John Wick style Inn was a fun concept that I enjoyed reading about.
There are a lot of similarities to this and FBAA and I would have liked to have it be a little more different, but I'm hoping book two will have the story turn into something of its own. Overall I enjoyed reading this story and I'm looking forward to reading book two especially after that ending.
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Reviewed in the United States on May 27, 2023
★★★★★ 5
Fallen Angels, fae, vampires, oh my!
Format: Kindle
Rating: 4.5 | Spice: 2 (but a good slow-burn)
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Main Characters: Huntyr and Wolf
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I couldn’t wait to read this book; there was so much hype about it! And there was no doubt why. I fell in love with the characters and the plot itself. This book is mainly plot driven more than friction driven but it’s easy to follow along with. The characters are fun, easily understood. The main setting is at an academy where both the main characters are going through trials and building strength for the final test, The Transcendent. There are fantastic side characters as well. I loved the camaraderie between Huntyr and her friends. But we don’t like Lanson. 😆
We do have some plot twists that come into play throughout the book. Secrets and betrayal to be seen. I did adore Wolf and Huntyr’s relationship. It was a classic slow burn trope. They didn’t hit it off fast, but in time their feelings grew. I loved their banter, so sexy. Wolf is your next book boyfriend; Huntyr is your next vampire assassin independent bad-a*s female. Themes include loyalty, trust, self-discovery, a true slow burn romance. Side note: book ends on a angsty cliffhanger!
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Emily, thank you for writing this awesome novel and I cannot wait to devour Book 2, Blood So Brutal! 😍
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Happy reading, my lovelies! xo
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Reviewed in the United States on January 21, 2024
★★★★★ 4
“My heart bows to you and you only, Huntress.”
Format: Kindle
3.5 🌟
This book popped up in my KU recommended reading suggestions and the synopsis sounded like what I was in the mood for. I'm so glad I took a chance on it. I went into this knowing absolutely nothing about it and ended up really liking it. I love when this happens.
The main characters are likeable and I easily found myself rooting for them. There is a mystery element to each of their backstories that I enjoyed watching unfold and can't wait to get more of. Wolf, in particular, has me fixated. Love him.
I found this to be an entertaining, addictive read with a plot that moves along at a good pace. It reads so easily I found myself very reluctant to put it down. Lots of twists and turns and the angst is there. A good set up for the next book to come, for sure.
My issues with this book....the dialogue feels a bit juvenile at times and there is a repetitive over use of a particular word phrasing that I found myself giving the ole eye-roll to. There are, without a doubt, some pretty cliche moments that gave me a bit of the cringe. I think this could've certainly 100% benefited from more depth regarding the world building. Perhaps the world building was sacrificed to keep the pacing quick? Just a guess. Also, the lack of consistency of character for the FMC was really evident and so she feels quite illogical at times.
Overall, this was a fun and enjoyable read that hit the spot well enough for me. That ending certainly has me impatiently pining for book 2!
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Reviewed in the United States on June 18, 2024