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Description
Human C3a 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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| Sensitivity | 0.053 ng/mL | |||||||||||||||||||||||||||||||||
| 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 Complement Component 3a (C3a) 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 Complement Component 3a (C3a) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Human | |||||||||||||||||||||||||||||||||
| Synonym | Human Complement Component 3a ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Complement fragment 3a (C3a) is one of the proteins formed by cleavage of complement component 3; the other is C3b. C3a is a passive 77-residue toxin that binds to the C3a receptor (C3aR), a class A G protein-coupled receptor. It plays a major role in immune responses. C3a molecules induce responses through the GPCR C3a receptor. Like other aprotinins, C3a is regulated by cleavage of the arginine at its carboxyl terminus, generating a molecule with less inflammatory activity (C3a desarg). It is an effector of the complement system with a range of functions, including T cell activation and survival, stimulation of angiogenesis, chemotaxis, mast cell degranulation, and macrophage activation. It has been shown to have both pro-inflammatory and anti-inflammatory effects, with its activity counteracting the pro-inflammatory effects of C5a. | |||||||||||||||||||||||||||||||||
| 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 7 reviews
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Product Reviews
★★★★★ 5
A Vivid Account of Gallant Thru-hikers
Format: Kindle
This book a little confusing at first since he is telling the story of several hikers at once. So keeping them all straight takes some effort. The actual description of the hike, the challenges and all the people along the way does make for fascinating reading. I personally, have great respect for those who attempt this formidable hike. They truly deserve our respect. I don't think you could be disappointed reading this book.
WAS THIS REVIEW HELPFUL?YesReportShare
Reviewed in the United States on May 20, 2026
★★★★★ 5
An epic adventure with a rich cast of characters
Format: Paperback
Scout paints the Mojave as bursting with life, a poetic world full of chaparral and chinquapin, of creosote scrub and conifers. These wild vivid landscapes are a backdrop to the real heart of the story, which, like all great stories, is always about People.
The Human Drama, sometimes of People vs the Environment, but more often of People vs Themselves. Overcoming our own fears or injuries or burdens that life inevitably deals to us all. This book captures the pursuit of people stretching themselves, challenging themselves to change and grow.
He does an excellent job of capturing the "esprit de corps" of the trail, of near strangers and fast friends looking out for one another, and the crescendoing intensity as they race against Father Winter. We meet a wide ensemble of characters, which is one of the most magical aspects of any long hike. It is the great equalizer, where people from all walks of life come together and share fellowship.
There are numerous role models within this book. It is impossible to not be inspired by Jan, the quintessential Fixer, or the protagonists who demonstrate exemplary examples of forgiveness and reconciliation, and other humble benefactors and do-gooders who understand that their gifts are more than simply money. They are giving opportunity. The fixers, the reconciliers, the benefactors....we should all aspire to be these types of people.
I hope this is but the first of his trail books. I'm certain there are many of us who would enjoy reading about his adventures from New Mexico to Montana as well. How fortunate was he to meet such wonderful people, and how fortunate are we to share their experiences from the comfort of our homes.
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Reviewed in the United States on August 11, 2020
★★★★★ 4
OK book about hiking the Pacific Crest Trail.
Format: Paperback
I love books about people hiking the national trails. I thought this book was about the author's thru hike on the trail with his wife. Actually, this book had little to do with the Pacific Crest Trail and was mostly about the other hikers, their relationships, etc. It read like a fiction book, as the author often gave examples of conversations the other hikers had with other people, even when he wasn't around to hear them. He also gave examples of what they were thinking and even what the other hikers friends were thinking. There was no evidence of interviews, etc.
It is a very well written book, so I gave it four stars. If you like books about the relationships of thru hikers, especially their romances, I would recommend this book. But it reads more like a fiction book, in my opinion.
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Reviewed in the United States on April 2, 2026
★★★★★ 5
Walking from Mexico to Canada with a great cast of characters
Format: Paperback
I thoroughly enjoyed reading Journeys North. The story is about the Pacific Crest Trail and the people who hike it (thru-hikers) attempting to walk from Mexico to Canada. The author brings the trail to life across the story, but what really stands out is the story of the characters. You find yourself rooting for them as they encounter all kinds of challenges, and you get to know them through their backstory and past experiences that are shared. People attempt a thru-hike for many different reasons, and the story is enriched by their personal stories and histories that they carry with them each step of the way. Even their “trail names” had stories that often highlighted something about the characters themselves. One of the unique things about the book is the Pacific Crest Trail itself, as the author’s descriptions allow you to imagine what it looked like and felt like (including the aches, pains, and injuries along the way). The trail is ever present across this story of the people who walked it, bring them together, and then pulling them away from one other, sometimes for weeks or even months. The trail and mother nature create for an everchanging backdrop as the characters navigate their way north. I am not a thru-hiker and I doubt I would ever attempt to walk from Mexico to Canada, but I enjoyed tagging along on the adventure of Scout, Frodo, Blazer, Dalton, Tony, Nadine and many others whose lives intersected over the course of many miles. I found myself having a hard time putting the book down, particularly as the hikers neared the Canadian border. If you are looking for an armchair adventure along the PCT, or wondered what it would be like to tackle a thru-hike of a long trail, or are curious about the people who undertake such a challenge, you will enjoy Journeys North! I would imagine that if you are a PCT hiker you would find the book even more meaningful.
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Reviewed in the United States on August 4, 2020
★★★★★ 5
Incredible Life experience while hiking the PCT
Format: Paperback
As I began experiencing great stories about the "Camino" pilgrimage in Spain, books about "thru hikes" in the US seemed to find me too i.e. The Pacific Crest Trail, and did grab my attention for various reasons...
"Journeys North"-- To me, this story makes it more than apparent, that if you want to learn about yourself and open your "vault doors" so to speak, and free yourself of personal challenges-hiking the PCT could help. This book on it's own, is a wonderful treatise about the PCT, and taught me much about being open and honest with others to include oneself, and about being grateful."
This story "rings" of Freedom, is intense, and at times, raw. From the comfort of my living room... Frodo, Scout, Blazer, and company drug this old backpacker (me) a.k.a. "LightningLayne" along on a fantastic journey north clear from Mexico's northern border to Canada's southern one. Along the way, I noticed how the "goodness" in people replaces just about every negative the world throws in your face. Finally, I laughed, I cried, and even felt sorry at times for all the "participants." In the end, I was "grateful" for the individual and collective stories shared. This story is one that has burrowed in deep and is one I will never forget.
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Reviewed in the United States on February 2, 2021
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