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Description
Human DDC 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 handling and requirements: The detection range of the kit is not equivalent to the concentration range of the analyte in the sample. Before the experiment, it is recommended to estimate the analyte concentration in the
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 handling and requirements: The detection range of the kit is not equivalent to the concentration range of the analyte in the sample. Before the experiment, it is recommended to estimate the analyte concentration in the sample based on relevant literature and conduct preliminary experiments to determine the actual concentration in the sample. If the analyte concentration in the sample is too high or too low, dilute or concentrate the sample appropriately. If the sample type is not listed in the instructions, it is recommended to conduct a preliminary experiment to verify the validity of the test. Serum: Collect whole blood in a serum separator tube and place it at room temperature for 2 hours or at 2-8°C overnight. Then centrifuge at 1000×g for 20 minutes and remove the supernatant. Alternatively, store the supernatant at -20°C or -80°C, but avoid repeated freezing and thawing. Plasma: Collect the sample using EDTA or heparin as an anticoagulant. Within 30 minutes of collection, centrifuge at 1000×g for 15 minutes at 2-8°C. Remove the supernatant and test it. Alternatively, store the supernatant at -20°C or -80°C, but avoid repeated freezing and thawing. Tissue homogenate: Rinse the tissue with pre-chilled PBS (0.01M, pH 7.4) to remove any residual blood (lysed red blood cells in the homogenate may affect the test results). Weigh the tissue and mince it. 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 or in a homogenizer. 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 and analyze it immediately, or store it at -20°C or -80°C, but avoid repeated freeze-thaw cycles. Cell lysis buffer: Gently wash adherent cells with ice-cold PBS, then trypsinize and harvest the cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be harvested directly by centrifugation. Wash the collected cells three times with pre-chilled PBS and resuspend in 150-200 μL of PBS per 1 × 10^6 cells (protease inhibitors are recommended; if the cell count is very low, the PBS volume can be reduced appropriately). Disrupt the cells by repeated freeze-thaw cycles or sonication. Centrifuge the extract at 1500 × g for 10 minutes at 2-8°C, and remove the supernatant for testing. Other biological samples: Centrifuge at 1000 × g for 20 minutes, and remove the supernatant for testing. Sample Appearance: The sample should be clear and transparent, and any suspended matter should be removed by centrifugation. Sample Storage: Samples collected for testing within one week can be stored at 4°C. If testing is not possible, aliquot the sample into single-use aliquots and freeze at -20°C (for testing within one month) or -80°C (for testing within six months). Avoid repeated freeze-thaw cycles. Hemolysis of the sample can affect the final test results, so hemolyzed samples are not suitable for this test. 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 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: Add 500 μL of universal diluent to each of seven EP tubes. Pipette 500 μL of the 10 ng/mL standard working solution into the first EP tube and mix thoroughly to make a 5 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, as shown in the figure below. 3. Prepare HRP antibody working solution: 15 minutes before use, centrifuge 100 μg of concentrated HRP antibody at 1000 × g for 1 minute. Dilute the 100 μg of concentrated HRP antibody to a 1 μg working concentration with universal diluent (e.g., 10 μL of concentrated solution + 990 μL of universal diluent). Prepare immediately before use. 4. Preparation of 1× Wash Buffer: Dissolve 10 mL of 20× Wash Buffer in 190 mL of distilled water (Concentrated Wash Buffer removed from the refrigerator may crystallize, which is normal. Allow to stand at room temperature until the crystals have completely dissolved before preparing the buffer). 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 them 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 sealant and incubate at 37°C for 60 minutes. (Recommendation: Dilute the samples to be tested at least 1-fold with Universal Diluent before adding them to the plate. This minimizes the impact of matrix effects on the test results. When calculating sample concentrations, multiply by the corresponding dilution factor. It is recommended to run replicates for all samples and standards.) 3. Wash: Discard the liquid and add 300 μL of 1x wash buffer to each well. Let stand for 1 minute, shake off the wash buffer, and pat dry on absorbent paper. Repeat this process three times (a microplate washer can also be used). 4. Add HRP detection antibody: After washing, add 100 μL of HRP detection antibody working solution directly to each well. Cover with a film sealer and incubate at 37°C for 60 minutes. 5. Wash: Discard the liquid and wash the plate five times according to the procedure in step 3. 6. Add substrate: Add 90 μL of substrate (TMB) to each well, cover with a film sealer, and incubate at 37°C in the dark for 15 minutes. 7. Add 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 450 nm. Calculation of Experimental Results: Result Interpretation: 1. Calculate the average OD value of the standard and sample replicates and subtract the OD value of the blank well as the correction value. 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 it appropriately and retest. Multiply the sample concentration by the corresponding dilution factor when calculating the sample concentration. |
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| Sensitivity | 0.06 ng/mL | ||||||||||||||||||||||||||||||
| Theory | This kit uses a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Sample, standard, and HRP-labeled detection antibody are sequentially added to microwells pre-coated with human DOPA decarboxylase (DDC) capture antibodies. After incubation and washing, the sample is developed with the substrate TMB. TMB converts to blue under the catalysis of HRP peroxidase and to yellow under the action of acid. The intensity of the color is positively correlated with the amount of human DOPA decarboxylase (DDC) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | ||||||||||||||||||||||||||||||
| Source | Human | ||||||||||||||||||||||||||||||
| Synonym | Human Dopa Decarboxylase ELISA Kit | ||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | ||||||||||||||||||||||||||||||
| Composition |
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| Background | Aromatic L-amino acid decarboxylase (AADC or AAAD), also known as dopa decarboxylase (DDC), tryptophan decarboxylase, and 5-hydroxytryptophan decarboxylase, is a lytic enzyme. This enzyme uses pyridoxine phosphate (PLP), the active form of vitamin B6, as a cofactor. PLP is essential for the decarboxylation mechanism of AADC. In the active enzyme, PLP binds to lysine-303 of AADC as a Schiff base. | ||||||||||||||||||||||||||||||
| 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 | 0.15-10 ng/mL | ||||||||||||||||||||||||||||||
| Applications | Serum, plasma, 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
😍😍 BEST DEBUT NOVEL EVER
Format: Paperback
I’m having trouble finding accurate words to describe the way this book made me feel, but I am going to do my best.
To start off with basic elements, the character and world building are phenomenal. I feel a strong bond to not only the two main characters, Ara and Rogue, but to each and every character introduced throughout the book. The author did a stellar job of giving each of them unique personhood. All of the scenes are beautifully described. So much so that throughout the entirety of the book, I could see every scene: the towns, the castles, the meadows, the landscape. I have had difficulty with this and with distinguishing between outlying characters while reading in the past, but I did not have to think to remember details of world or character building because they flowed naturally within the story and were described well. I have read book series before that made me want to be a part of that world, but I actually felt like I got to step into Auryna and Ravaryn!
The plot twists!! Although this is not a suspense novel, it still had me on a rollercoaster of emotions and on the edge of my seat from beginning to end. I haven’t cried actual tears over a book since I was in high school (and I’ve read a LOT). This book finally broke the floodgates in the final few chapters. Multiple times. And we love a good cliffhanger. It truly made me FEEL.
THE SPICE is a solid 3.5/5. Some of the scenes had me flushed, some had me taking notes, some just had my jaw slack and my mouth hanging open. Bravo, JD Linton, bravo.
The relationships: friendships, family, romantic, ALL of the relationships in this book have so much meaning. The author does a great job at making you feel the love, the anger, the peace, the frustrations, the safety, the familiarity, etc. between the characters.
Ara and Rogue. I can not say enough and I also do not want to say too much. Just know that I feel like I know them both, to their core. I know what their childhood looks likes, their darkest moments, their biggest fears, their dreams and passions, what they want in life… The POV switches were seamless. I am so happy this author decided to let us see from both sets of eyes.
I can not wait for book two after that cliffhanger. And there is SO much potential for at least one prequel, I can’t wait to see where this author goes! I hope this series continues and flourishes. Fingers crossed!
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Reviewed in the United States on December 5, 2022
★★★★★ 5
Amazing!
Format: Kindle, Format: Kindle
This book was phenomenal, I devoured it within a few days! For this being a debut novel, it is fantastic and I would’ve thought the author was a seasoned author. I have zero complaints about this book.
Let me start by saying that the world building was phenomenal. I could picture everything in my head because of how detailed it was — that’s how good it was written. And I absolutely love the “captive/captor” trope so much, it’s become one of my favorite tropes, so I was pleasantly surprised to see that this book had that.
I loved the banter between Rogue and Ara — they’re both snarky and witty, plus with the romantic tension, it made the dialogue that much better. Speaking of romantic tension, yes there is spice but not so much of it that it overrides the plot, which I loved. For me, this would probably be on the 3/5 level of spice.
This book had a ton of plot twists and I thoroughly enjoyed it.
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Reviewed in the United States on November 13, 2024
★★★★★ 4
High on Tropes and Satisfaction
Format: Kindle
This is a great Romantasy book full of action, adventure, and everything you look for in this genre.
I won’t lie: it does kinda feel like the author found every common trope from every successful book of this kind and threw them all into this novel. But if it ain’t broke, don’t fix it. Especially in romance, there’s a large audience who has specific expectations, and they want them every time. Nothing wrong with that and many times I’m one of them.
I have no idea what defines a spoiler honestly, so
spoiler alert!!!!!!!
Tropes include:
Only one bed at the inn/bar
Dissatisfaction with life before hunk appears
Lost royalty
The chosen one
Montage of dress up time followed by shocked hunk
Forbidden romance between two from rival peoples
Power that cannot be controlled, simply guided/asked
Gathering intel at the inn/bar
FMC who knows how to fight/use weapons well
There’s probably more but no need to list them all.
Good story and I would recommend!
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Reviewed in the United States on June 14, 2024
★★★★★ 5
AMAZING debut novel!!!
Format: Kindle
Plot ⭐️⭐️⭐️⭐️
Spice 🌶️🌶️.5
Romance 💘💘💘
Vibes ⭐️⭐️⭐️⭐️⭐️
Dual 1st person POV - Ara (26) & Rogue (39 - but looks mid-20s: they can live hundreds of years so this isn't that large of a gap as it could've been which I heavily appreciate lol)
Tropes: enemies to lovers, fae/human wars (deep hatred for each other), shifters (dragons- MMC can only partial shift with wings), one horse, one bed, touch her and d!e, found family, abduction turned to freedom
The Last Storm is the debut novel from JD Linton and let me tell you, you guys NEED to read this. The plot was engaging and the editing was was amazing (especially for a debut novel). Our FMC, Ara, is stuck in her gilded cage longing for a life outside of her small town. She uses her books to escape and live vicariously through the pages (honestly, relatable). After her father announces her betrothal to her childhood friend (to whom she has no romantic feelings for), Ara tumbles unknowingly into a desperate plot trying to stop the humans from slaughtering the Fae.
As one can expect from an enemies to lovers / kidnapper/captive romance, Ara fights her attraction and lust towards our MMC, Rogue (the King of the Fae), for as long as she can. Upon seeing Ara for the first time, Rogue is instantly aware that she is his fated mate (not a spoiler). Since she is the General's only daughter, he plans to abduct her and use her as leverage to stop the brutality. During Ara's time in Rogue's captivity, their banter and chemistry continue to rise until they finally boil over and come together (quite literally, and many times I may add 😉).
Here's what I LOVED:
- Rogue continuously seeks advice from his elders and deeply respects their opinions and life experience and tries to implement their recommendations
- Rogue makes many mistakes in the beginning but we see him actively work on not repeating them as the book progresses. The level of self-awareness and his ability to change his behavior was impressive
- The magic system is intricate and we have only scraped the surface. As the series continues and Ara progresses in her powers, I'm sure we'll get to see more of this. I absolutely LOVE the messaging system that is used in this book.
- Ara's struggles are so human and so raw. She is experiencing so much guilt and pain and hurt and getting to see her work through each of these emotions is inspiring. Especially as her and Rogue get closer and she learns she can lean on him as well, that she is not alone.
- While this is the start of a series, there is NO cliffhanger! There's a bit of a teaser of something major that is going to happen at the start of the next book, but it's not a cliffhanger in the sense that we aren't sure if someone is going to live or d!e or if they'll be separated. For that, I am very thankful! This book was so much fun that I will definitely be returning to book 2, even if it takes several months (or longer since this is an debut author) to publish!
- Lastly, the cover is GORGEOUS! And I love the title!
I'll copy a few of my favorite quotes below so you can have a little taste of the author's writing and the world she's cultivated. 😊
Top Highlights from The Last Storm
On days like this, when my heart was heavy and my mind clouded, I resorted to books— to escape, to forget, to find freedom where I had none.
If I were to marry him, my face would always be turned to the window, searching for more, and if not that, I would be a shell of the person I am now.
I stepped back to admire her, thr0bbing at the sight. She was the most beautiful woman I had ever seen. To ever exist. Nothing, no one, had ever deserved to be worshiped more. All men should be made to kneel before her. But she would have to settle for me.
The taste of her met my t0ngue as my scent merged with hers, forever branding her. Mine. I l!cked the wound. Hers. Completely and utterly hers.
I didn’t claim her in ownership. I claimed her as my one. Devoted myself to one. With that mark, my body and soul were bound to her. I would never be with anyone else, emotionally or physically. It would be her or no one, until my last breath.
“Scream my name. Let everyone know who I belong to.”
I had never really cared about the weather before, but now, clear skies meant everything to me, and I was grateful to see another calm morning.
“There will never be another woman for me.” He paused. “Ever.” I stilled at his words. “What… Why?” “This”— his thumb slid down across the mark—“ is a symbol of… surrender. I know you believe that it was my claim upon you, but it wasn’t. It never was. I bound my body and soul to you, little storm.”
“I also know that it is more than this tiny, insignificant mark on your skin that binds me to you. It’s you. All of you. Your strength and resilience. Your determination to endure no matter what fate throws at you. Your love for love and stories and hope. You are entirely the opposite of everything that I am and I would gladly wear your shackles if it meant I could have you.”
My mate. Mine. And then everything shifted and I understood. I understood everything. The surrender. The deep, soul-craving longing. Bound. I was bound to him. Body and soul. Entirely his. “I would’ve waited forever,” he whispered back, understanding.
Seriously, everyone.. add this to your TBR!!
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Reviewed in the United States on November 14, 2022
★★★★★ 4
A Decent Fae Romance
Format: Kindle
** 3.5 Stars rounded up to 4 **
First off I want to say that I found the author, J. D. Linton, on TikTok right before the release of the sequel. I bought the first 2 books to support her release, so congrats to her for the release of the sequel!
Overall, I liked the story & plot, I liked the characters, and I liked the spice. The downfalls: not enough development between the characters for the romance, and it leaned a little heavy into the tropes. A lot of stuff is told to you, which could have been used as devices to drive the plot and emotions of the characters.
🛑🛑 Mild Spoilers ahead, I tried to hide the obvious ones 🛑🛑
This is very obviously a fated mates, which is a trope I usually really enjoy. But I wish the relationship between Ara & Rogue had much more development before it's realized that they're mates, instead of Rogue knowing almost immediately and using it for nefarious purposes (at least in the beginning before they get to know each other). By revealing this within the first quarter of the book, I feel like it leaves less room for them to fall for each other organically (albeit with help from the mating bond) and they love each other because of the mating bond. I was disappointed as soon as Rogue know (literally only 5% in) and I literally made a note: "As much as II love a good mating trope, I wish we had to work for it a little more. Where's the fun in just telling us?" I believe that by holding out and feeding the reader snippets of a potential bond, it would've been more rewarding as a reader.
I also wish there was more world building - we are told of a war between human & fae but don't get to really experience any of it. Ara is sheltered in her human home, then sheltered in Rogue's castle. There's bits and pieces about what the war has done on either side - but we're more told of the aftermath and don't really experience any of it. Ara's father is supposed to be the king's #1 general - yet he is at home with his family & with Ara for the first couple chapters. Her, her family, nor her village seem to be affected by the 10 year war going on on their borders. I wish there was a little more setup to make this conflict - an actual war - feel more than a skirmish between fighting territories.
Linton could also be a little repetitive - with the biggest culprit being when Ara is upset she "brings [her] knees to [her] chest]" and either sits like that or cries. Every time she is upset this phrase is mentioned. I would get it if this was her crutch, or how she copes with grief and stress, but that should be explained why she does it so often or it becomes repetitive. I started to get annoyed with how often she would sit like this solely because it happens every couple chapters.
However, I did really like the spice. I love an enemies to lovers trope, especially when it results in spicy scenes. The spicy scenes weren't anything new, but they were fun. Wish there were more but that could also just be me - there is no such thing as too much spice 😂
Would I read again? Probably not, I'm super picky with rereads. Although I did genuinely enjoy my first read through!
Will I continue the series? Probably, at least for the sequel. As for #3, kind of depends on where I am with my TBR once that is released
All in all an enjoyable, fast paced read
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Reviewed in the United States on March 8, 2024