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Description
Rat TIMP-2 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: 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: 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. Plasma: Collect the specimen using EDTA or heparin as an anticoagulant. Centrifuge the specimen at 1000 × g for 15 minutes at 2-8°C within 30 minutes of collection. The supernatant can be assayed or stored at -20°C or -80°C, but avoid repeated freezing and thawing. 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 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., 1 g of tissue sample to 9 mL 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 the supernatant can be assayed. Cell Lysis Buffer: Gently wash adherent cells with ice-cold PBS, then trypsinize and collect cells by centrifugation at 1000×g for 5 minutes. Suspension cells can be collected directly by centrifugation. Wash collected cells three times with ice-cold PBS and resuspend 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 freeze-thaw cycles or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, 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, avoiding repeated freeze-thaw cycles. 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 enzyme conjugate are sequentially added to microwells pre-coated with a Tissue Inhibitors of Metalloproteinase 2 (TIMP-2) 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 Tissue Inhibitors of Metalloproteinase 2 (TIMP-2) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Tissue Inhibitors Of Metalloproteinase 2 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Tissue inhibitor of metalloproteinases 2 (TIMP2) is a gene and corresponding protein. This gene is a member of the TIMP gene family. This protein is considered a metastasis suppressor. It is a specific inhibitor of MMP-2, binding to activated or inactive MMP-2, terminating MMP-2 activity and effectively inhibiting MMP-2's collagen and gelatin degradation activities, thereby suppressing cancer cell metastasis and invasion. Numerous studies have shown that TIMP2 and TIMP-2 are two indicators of tumor invasiveness, playing a crucial role in the metabolic balance between extracellular matrix synthesis and degradation. Disruption of the balance between MMP-2 and TIMP-2 in tumors is a key factor in tumor progression. | |||||||||||||||||||||||||||||||||
| 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 | Serum, plasma, tissue homogenate, cell lysate, cell culture supernatant and other biological fluids |
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4.6 ★★★★★
Based on 20 reviews
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Product Reviews
★★★★★ 4
Oh this was adorable and cozy and spicy
Format: Kindle
Oh this was adorable and cozy and spicy and I had so much fun with this book!
The relationship between Cammie, Finn and Reid is good. I love how doting both men are of her and her kids, always providing them with a safe space to be themselves. They were patient with Ben and I loved seeing that represented in this book. The plot was done well and it was well paced with quiet, family moments as well as drama to keep us on our toes.
I do wish we'd gotten more moments just between Finn and Reid to showcase their strong relationship (they've been together for two decades!) and while we got something, both men were first and foremost focused on Cammie. It makes sense why this was, there was a lot of hurt that needed to be soothed and I'm glad she is treated like the queen she is, but I did yearn for more of the alpha-omega couple.
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Reviewed in the United States on September 29, 2025
★★★★★ 5
A Beta’s Dream!
Format: Kindle
I loved this book!!
Unwanted follows Cammie, a single mom and Beta, working two jobs, trying her best to raise a son and daughter since her a$$hole ex Alpha kicked them out after meeting his scent match.
Being a Beta from the wrong side of the tracks, Cammie is often the victim of rumors and blatant disrespect from several alphas in town; and while in a panic searching for her daughter who somehow managed to wander outside of the home, she finds herself in a panic when she comes across a police officer. Rather than help her, Cammie finds herself feeling threatened and helpless against the alpha’s behavior.
That’s when Reid, one of her Alpha neighbors (and super hot firefighter) swoops in to her rescue. Sparks fly, but Cammie thinks nothing of it since she is so used to being disregarded. Little does she know, Reid goes home to his Omega, the adorable and caring Finn, and confesses that he thinks he’s met their match.
What follows is a beautiful hurt/comfort romance, with lots of steam, and found family perfection! Reid and Finn worship Cammie and absolutely adore her kids. They’ll do anything to protect them and cherish them, to erase all the hurt and rejection that their bio dad put them through. It’s such a heart warming and emotional story—perfect for when you’re feeling like something low angst with princess treatment for not just Cammie, but her kids as well.
I definitely recommend picking this one up!!
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on June 28, 2024
★★★★★ 5
Emotional read with children
Format: Kindle
Cammie is a single mother of two children whose ex-boyfriend, who is an alpha that he met his scent matched omega and dumped her via messenger on his phone then the next day put their house up for sale. Cammie moves back to her hometown, but because she has no support, is working two jobs to keep her and the children in a small rented house. One night her four year gets out of the house and she meets a neighbour, but she refuses to make the same mistake again by getting involved with another alpha, but Reid is convinced they are scent matched mates even though Cammie is a beta. Reid and his omega Fin start to court Cammie and her two children that they belong together as a pack and family. Cammie's ten year old son is a tough one to convince, but the scenes between him and Reid and Fin, they were so good. I was tearing up at times, found family was a big theme and I really liked this book. I highly recommend that you try it too.
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Reviewed in the United States on February 16, 2025
★★★★★ 5
It’s an experience
Format: Paperback
⭐⭐⭐⭐⭐
**A Riveting and Emotionally Charged Journey**
"Unwanted" by Sirena Song is an absolute masterpiece that grips you from the very first page and doesn't let go. The storytelling is impeccable, weaving together a tapestry of raw emotion, suspense, and heart-wrenching beauty. Song's ability to delve deep into the psyche of her characters makes them come alive, making you feel their pain, joy, and struggles as if they were your own.
The plot is brilliantly crafted, full of unexpected twists and turns that keep you on the edge of your seat. Each chapter unfolds like a piece of a complex puzzle, gradually revealing the depth of the narrative. The themes of love, rejection, and redemption are explored with such nuance and sensitivity that it leaves a lasting impact.
Sirena Song's prose is poetic and evocative, painting vivid pictures with her words and creating an immersive reading experience. The character development is outstanding, making it easy to become emotionally invested in their journeys. The ending is both satisfying and thought-provoking, leaving you pondering long after you've turned the last page.
"Unwanted" is not just a book; it's an experience that touches the soul. I highly recommend it to anyone looking for a powerful and unforgettable read. Sirena Song has truly outdone herself with this one.
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Reviewed in the United States on June 30, 2024
★★★★★ 4
Totally Sweet
Format: Kindle
🔥OmegaVerse
🔥Single Mom
🔥Scent Match
🔥Emotional Trauma
Well written, emotional omegaverse about a single mom of 2 with an ex-Alpha boyfriend that found his Omega and ditched his family. Cammie has a meet cute with Reid and Finton separately but they both find her to be their scent match.
This book is totally sweet with lots of drama but lots of proving to Cammie that she is loved and wanted.
My Personal Ratings:
5 Stars: Loved it! Couldn’t put it down or obsessed about it while needing to get to the end of it. Had to share some bits with my hubby!! Usually read in less than 2 days cause a girl has got to sleep sometime. Will be reading everything this author has to offer and most likely reread the series as next books in the series are released.
4 Stars: Really Good. Whether the story drew me in or a specific character, will definitely continue the series and look into more books by this author.
3 Stars: Enjoyed something about the book, whether it be the storyline or a character but also may have not liked something. Sometimes the book is great but needs some TLC from an editor. May or may not continue the series.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on July 5, 2024
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