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Description
Rat ESM1 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 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. 3. 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. 4. 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. 5. 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. 6. 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 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: Take seven EP tubes and add 500uL of universal diluent to each. Pipette 500uL of the 10ng/mL standard working solution into the first EP tube and mix thoroughly to make a 5ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube serves 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 Endothelial Cell Specific Molecule 1 (ESM1) 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 Endothelial Cell Specific Molecule 1 (ESM1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Endothelial Cell Specific Molecule 1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Endocan, formerly known as endothelial cell-specific molecule-1, is encoded by the ESM1 gene and is a 50 kDa soluble proteoglycan composed of a 165-amino acid mature polypeptide and a sulfated dermal chain covalently linked to a serine residue at position 137. The expression of this gene is regulated by cytokines, suggesting a possible role in endothelial cell-dependent pathologies. Studies have shown that endocan expression is increased in the presence of pro-angiogenic growth factors such as vascular endothelial growth factor (VEGF) or fibroblast growth factor 2 (FGF-2). Endocan overexpression has been detected in highly vascularized cancers by immunohistochemistry using a monoclonal antibody against endocan/ESM-1. | |||||||||||||||||||||||||||||||||
| 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 | 0.156-10 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, tissue homogenate, cell lysate, cell culture supernatant and other biological fluids |
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4.3 ★★★★★
Based on 20 reviews
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Product Reviews
★★★★★ 5
I have a few instructional writing books I reread; this is one of them.
Format: Paperback
Plot & Structure: Techniques and Exercises for Crafting a Plot That Grips Readers from Start to Finish by James Scott Bell may be the best book for plotting and creating structure in fiction stories that I’ve read.
I have a few instructional writing books I reread; this is one of them.
Plot & Structure works for writers who don’t use outlines and for those who do. Throughout Bell acknowledges and addresses how his ideas about plot and structure can be used by all writers. This isn’t a “my way or the highway” kind of book. Bell gives practical advice for all writers.
What’s so great about Plot & Structure? The biggest plus is Bell’s practical and down to earth strategies and clear explanations.
Here are the other greats:
Bell’s approach isn’t a how-to formula; it’s more of a guide. His genre is thrillers, but he addresses the needs of other genres as well as those of literary fiction.
He provides many strategies writers can use to approach plot and structure; plus, he encourages readers to find their own way of working.
The exercises at the end of the chapters are practical. If you try some of them, you’ll discover that they aren’t busywork but helpful approaches to solving writing issues.
Unless you’re a pro with so many publications behind you that you don’t need help, this isn’t a book to read once and put away. It’s a book you’ll come back to and get more out of with a second or third reading.
If you want a formula or step-by-step program, this isn’t a book for you. He gives choices so you can find you own style and way of working. Which is one of the things like I about the book. It’s like going to a writer buffet and choose the strategies that work for you.
If you are someone who writes without an outline, you are going to be surprised because Bell doesn’t leave you out of any of the equations. He has strategies for all writers.
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Reviewed in the United States on November 13, 2014
★★★★★ 5
Great advice, guidelines, and suggestions: Bell writes with a contagiously inspiring attitude
Format: Paperback
I admire good organization when it comes to most things, especially in a book that purports to teach about structure. The organization of the book makes it easy to go back and reread a part; for example, if you are working on a beginning, middle, or end section in your fictional story, there are corresponding chapters for what you need. Each chapter has ending exercises that I imagine to be helpful, but I haven't actually worked them through although I've read them.
You will read about elements that you need to know to how to do to write an appealing story, like characterization, settings, dialogue, and scene selection. It is recommended that you get a book with these elements as the main focus for a more detailed, expansive treatment (if you haven't done so already). Bell is most helpful on how to construct a scene. However, he does not have much to say about description, other than that it has a tendency to slow the plot and you should give details as needed but not more than that (on the line of a need-to-know-type of basis).
Bell also has sections on how to brainstorm plot ideas and test whether the ideas are worthy of attention. This was something I needed to do with my last story idea that burned and crashed. Furthermore he tells of "plotting systems" used by many writers, which are the-strict-only-by-the-outline group (OPs) or the free-rein-no-outline group (NOPs) and writers using a little of both.
The suggestions and tips are conveyed in a way that you can take and adapt them to your style. The chapter on common plot problems was very helpful. He even tells of how he squeezes in his writing time. Bell's way doesn't work for me, but you can take it and suit it to your own circumstances.
What I like about Bell is that he writes in a non-demeaning way to new writers. There are some writing books which take punches at new writers, be it ever so subtly or blatantly (I would rather not name any names), even though these books may contain invaluable writing advice and tools. It is just refreshing to have Bell's attitude. He motivates me to write and be true to myself, especially in his introduction. So far it is one of the books I keep going back and rereading parts that are pertinent to what I am writing.
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Reviewed in the United States on February 8, 2006
★★★★★ 5
As Much About Life As Writing
Format: Paperback
Anne Lamott is not a cheerleader, more like the Burgess Meredith with the water bottle and bucket in Rocky's corner between rounds -- I'm also guessing she wouldn't wilt if she had to slash your eye open if like Rocky it got sealed shut. She knows you are going to get hit hard, and she reminds you that you know it too. She tells you not to get distracted by that which doesn't matter to the process of writing. Much of this she learned from her father, who was also a career writer. He taught her it was the doing that mattered, not the surrounding mechanical functions that seem like they matter.
What struck me repeatedly in Lamott's mini-lessons was her deep understanding of process -- that output of a work is not so much the full work itself, but an assembly of building blocks, one at a time, each a commitment, and only in totality something more. She does not advocate bonehead process or ridiculous formulaic mandate - this is not a how-to manual -- she just wants us to care about what we are doing and accomplish it in a series of heartfelt steps. There are no shortcuts, it's a little more each day, a continuum that adds up to a satisfying and cohesive whole. This is not breakthrough thinking, but it's a lesson we need to learn over and over, and it's not just about writing. Creative process is the heart of innovation. Think of all the elements that make the iPad great. If all the elements weren't great, it would not be great. Same with a restaurant menu and wine list. Same with an office skyscraper or memorial monument. Same with a short story, same with a novel. Summary impression rests in the details, all the many tiny parts or moments -- and all those details require hard thought and careful design.
Lamott is smart about this, she tells you that getting it right is not going to happen out of the gate and unnerving strides at perfection can be your worst enemy. She has an excellent descriptor for the real quality of the first drafts to which we aspire. I'll let you discover that on your own so the word does not get scraped here. Her point is, just get the words out, work on making them better later, a layer at a time.
She also allows us not to obsess unnecessarily with locking the full road map before we explore, because again that can impede our work. How far do we need to see ahead? "About two or three feet ahead of you" is plenty she tell us, quoting E.L. Doctorow: "..writing a novel is like driving a car at night. You can only see as far as your headlights, but you can make the whole trip that way." She says this is "right up there with the best advice about writing, or life, I have ever heard." I tend to agree.
There is tremendous empathy in Lamott's world view, she offers a sense of shared experience that is reinforcing and comforting. Lamott talks about the imaginary radio station playing in your head -- another colorful descriptor I will let you discover -- that tells us over and over again why we can't do something, why the work we are doing is neither good nor worth doing. Learning to turn off that radio is our key to moving forward, we all hear it from time to time, but when it becomes perpetual, that is when our ability to create interesting work stops completely.
Lamott is just so honest and clear about all the factors that stop us from moving forward because she not only has experienced them, she continues to experience them. She does not position herself as a guru or weekend seminar success evangelist, but simply as someone who can reflect on problems of creativity because she deals with problems of creativity endlessly in her own life. She is even more honest in telling us that no one can make these problems go away once and for all, certainly not with any form of temporal success. All we can do is know that these obstructions will always be there, so we must embrace confronting them. Sometimes it really is good to know that none of us are experiencing roadblocks on our own, the fact that someone like Lamott tells you she is experiencing what you are experiencing is precisely the empathy that builds strength and resistance because the experiences are shared, bad and good. Her humility is reinforcing and refreshing and uncompromisingly inspiring.
"Bird by Bird" is not a long book, it can be read if you wish initially in a single sitting, but it is the kind of book you will find yourself coming back to for this chapter or that, this phrase or that. Lamott writes with good humor, even when she tackles very difficult and personal matters of her own life and those around her. The more I think about her framework, the more I am convinced it is much more broadly applicable then perhaps she even considered. I see the guidance as useful in company life, in financial life, in family life, in political life, and in government life. All of these require effective process to get them right, there are no shortcuts, and the rewards can be the smallest where the challenges are the greatest. That does not mean the rewards aren't meaningful, but it is the context of those rewards and the expectations that one sets for success that truly inform us when we are steering toward a final draft.
Review excerpted from my blog:
[...]
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Reviewed in the United States on October 3, 2011
★★★★★ 5
Attention: Writers! This book is for all of us, new or old pros at the craft.
Format: Paperback
Some - because it's too many to list ALL - of reasons I loved Bird by Bird as a writer, an entrepreneur and a human being with stories to share and thoughts to voice - just like you.
I wish I had read the Kindle version so I could highlight every chapter and re-read them later, but I got the physical book to add it to the small shelf of books I still keep in my office, and next to the copy of my most favorite book on the topic: On Writing by Stephen King. Bird by Bird is now the 2nd best and useful book I have read on writing.
If I weren't inspired to write before, I felt my body fill up with reasons that now draw me to writing. If I had voices of doubt, the ones that slip into your head at 4am and whisper, "Who are you to write? What do you have to say that others want to read?", which I did, I am now courageous enough to ignore them. They don't go away but I can say, screw you, I'm writing anyway.
I now can't not write and this feeling is one that I just can't put a price on. For that alone, I'm overjoyed. But there's more.
For starters, it's the way she tells you to write about your childhood. It's the simplicity of this advice, and the way she just calls you to do it. I loved her just for that. Write about your childhood even if it was terrible, even if it was dark and lonely, for it shaped you into who you are and who you are is a unique voice for the world to hear. Okay I added all of that but she says it more beautifully and the gist of it says that thou must write about thy childhood especially when you do not know where to start. Make it a good story, turn the bad characters into a description that if they were to read your book, they would not recognize themselves except for their actions - that's apparently how you save yourself from libel - and then just write.
I cried at various points in this book and the whole spiel about childhood was one of them, but the others were when she described why we write, and what brings us to the blank page, and how it's not about being published - and it really isn't, I've been published twice and my book has ranked among best-selling categories and sold thousands of copies and it was fun yes but writing is about so much more.
If you weren't enough before you published, she quotes someone, then you won't be enough after, and that will stay with me. We all write for our own reasons, and if you feel drawn to writing, if you feel a call to writing and you have been resisting it, stop. Ignoring this urge is like neglecting hunger or thirst just because you are too stubborn to accept the laws of nature. Go with the flow, drop your excuses and write what you feel called upon to write.
I also love the writing style of Bird by Bird. She does not break down writing into distinct categories and address each. She simply tells us stories and personal experience and her amazing nuggets of wisdom come through, just oozing out of the page. It is the stories that help you remember the bigger points she was making, and a very similar style as King's On Writing, which helps you learn not just about writing but about the writer's life and highs and lows and how writing integrates into their life, and the big picture.
Bird by Bird is sweet, refreshing, funny, and even if Lamott over-dramatizes the life of an author - or perhaps, mine is under-dramatic, who knows - I love her for it. I love that she was oh so vulnerable, and how she dished out tough love and great advice and in the end, simply encouraged us to write. Just write those stories down and do it for reasons that go well beyond publication, fortune, fame, or other dreams that you may have for your writing, because writing is its own sweet delicious fulfilling reward. Add this book to your list of must-read books, my writer friends, and let's put our stories out in the world if not for anyone but ourselves.
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Reviewed in the United States on March 28, 2015
★★★★★ 4
Lamott's "Bird by Bird" is a real tweet
Format: Paperback
In "Bird by Bird," Anne Lamott relates the growing pains felt by all fledgling writers, including herself at one time, and the importance of staying on course until the job of writing is done.
The book is sown with plenty of humorous anecdotes, zany analogies and colorful metaphors - in short, the emblematic, original style of writing for which Anne Lamott is best known.
"Bird by Bird" does not deviate from this signature style by drifting into territories of discussion about proper grammar, form and other pedestrian aspects of writing. She feathers her nest in a creative, engaging format, filling it with stories of her earlier days as a writer and interspersing them with tips and lessons learned along the way.
There is no elitism in "Bird by Bird." Lamott demonstrates her humanness in the incidents she shares. When a friend calls to say that her book has been published readers will empathize with Lamott's natural feelings of jealousy and inferiority. When she ends up writing about those unpleasant emotions, the lesson becomes apparent that any topic - especially one that is universally felt, experienced and, therefore, understood - has the potential to be fashioned into the written form.
Later on, the author reveals the anxiety and knuckle-cracking anticipation she experiences while awaiting feedback on a manuscript submission. It is impossible not to feel jitters of sympathy as we wait for the outcome to be revealed.
For the most part, "Bird by Bird" covers the emotional and creative expanse experienced by the writer from the moment he first coaxes his work onto paper until such time that he deems it finished. Lamott reminds writers that aspiring to have their works published should not trump the sheer joy of writing for writing's sake.
The book could prove a valuable addition for a writer who has already begun to see some positive affirmation and movement where his writing is concerned. For a beginner, however, this book might miss the mark due to its absence of writing fundamentals. Lamott does address this subject, however, in a general sense when she alludes to writers' groups and writers' workshops.
The few flutters of grandstanding that occasionally manifest as Lamott tells her story seem more than justified; she deserves recognition, to be sure. Despite a thorny start, Lamott has arrived - observing, reflecting and, finally, writing it all down from her position in the catbird seat.
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Reviewed in the United States on March 13, 2009
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