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Description
Rat C4 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 specimens using EDTA or heparin as anticoagulants and centrifuge them at 1000×g for 15 minutes at 2-8℃ within 30 minutes of collection. The supernatant can be tested or stored at -20℃ or -80℃, but repeated freezing and thawing should be avoided. 3. 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 400 ng/mL). Then dilute to the following concentrations: 400 ng/mL, 200 ng/mL, 100 ng/mL, 50 ng/mL, 25 ng/mL, 12.5 ng/mL, 6.25 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 400 ng/mL standard working solution into the first EP tube and mix thoroughly to make a 200 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 Complement Component 4 (C4) capture antibody. After incubation and washing, the assay 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 Complement Component 4 (C4) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Complement Component 4 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Complement protein 4 (C4) is a protein involved in the complex complement system, derived from the human leukocyte antigen (HLA) system. It plays a number of key roles in immunity, tolerance, and autoimmunity, along with numerous other components. Furthermore, it is a key element connecting the entire systemic recognition pathway, initiated by the antibody-antigen (Ab-Ag) complex, with other effector proteins of the innate immune response. For example, severe complement system dysfunction can lead to fatal illnesses and infections. However, C4 protein is derived from a simple, bifocal allelic model, the C4A-C4B gene, allowing for abundant variation in the levels of its respective proteins within the population. The C4A-C4B genetic model, originally defined in the context of the Chido/Rodgers blood group system, is currently under investigation for its possible role in the risk and development of schizophrenia. | |||||||||||||||||||||||||||||||||
| 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 | 6.25-400 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma, and other biological fluids |
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4.2 ★★★★★
Based on 604 reviews
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Product Reviews
★★★★★ 5
Sturdy aggressive chew toy.
Color: Green
Sturdy aggressive chewing dog toy. My 8 year old Rott loved it. Non-stop chewing for 1st day. Usually only buy her Kong toys, and based on reviews decided to try this one. Great product.
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Reviewed in the United States on May 21, 2026
★★★★★ 1
Not really indestructible if it’s plastic
Color: Green, Color: Green
We received this toy yesterday evening and in about 10 mins of giving it to my dog(Labrador) He was able to bite into the plastic parts. I had to take it away from him. So this product is not for aggressive chewers. In fact any dog small or big can bite into the plastic parts. Now when my dog plays with toys he focuses more on the ends of the toy if he deems it to be a chew toy. So he hasn’t gone to the squeaky part yet as I had to take it away from him.
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Reviewed in the United States on May 12, 2026
★★★★★ 4
Dearly departed
Color: Green, Color: Green
This toy lasted two sessions. Two. Sessions. I don't really fault the toy. I fault the dipstick in the donut suit.
Our dog did love this toy.
He did look extremely cute carrying a caterpillar around.
We did worry about property damage when he would joyfully toss the toy.
I find the claim of "indestructible" to be exaggerated.
We had some very tense minutes when we couldn't find the caterpillar's head. We kept looking at dog ingress points and dog eggress points with more and more worry. He did NOT swallow the head, it had rolled under a piece of furniture. He is blissfully unaware of how lucky he is.
If he was half his size, this would have been a marvelous toy.
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Reviewed in the United States on April 21, 2026
★★★★★ 5
Chew toy that holds up to aggressive chewers
Color: Green
Bought this for my 85 lb aggressive chewer. She loves this toy and gnaws on it constantly. My only complaint is that she likes to bring it to bed with her and I have to listen to "chomp, chomp, chomp" while falling asleep!
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Reviewed in the United States on May 20, 2026
★★★★★ 5
It lasts!!!!
Color: Green
My dog LOVES this toy! He's a very aggressive chewer and so far this is the only toy that has lasted him. My guess is that he will eventually destroy it, but not for a very long time. He chews it so much that it gets "spikey" from all the chewing, but so far it's the only toy I gave found that lasts him. I've replaced it once because it gets yucky looking after all the chewing but that's my choice. I definitely recommend thus one for an aggressive chewer!
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Reviewed in the United States on April 15, 2026