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Description
Rat CX3CR1 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 preparation and requirements: 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 and
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 preparation and requirements: 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 and mince the tissue. 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. 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 collect the supernatant for analysis. Cell Lysis Buffer: Adherent cells should be gently washed with pre-chilled PBS, then trypsinized and harvested by centrifugation at 1000×g for 5 minutes. Suspension cells can be harvested directly by centrifugation. Collected cells should be washed three times with pre-chilled PBS and resuspended 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 freezing and thawing or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and collect the supernatant for analysis. 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 utilizes 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 Chemokine C-X3-C-Motif Receptor 1 (CX3CR1) 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 Chemokine C-X3-C-Motif Receptor 1 (CX3CR1) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Chemokine C-X3-C-Motif Receptor 1 ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | CX3C chemokine receptor 1 (CX3CR1), also known as bifurcating protein receptor or G-protein-coupled receptor 13 (GPR13), is a protein encoded by the CX3CR1 gene. As its name suggests, this receptor binds to the chemokine CX3CL1 (also known as neuronal or bifurcating protein). Expression of this receptor appears to be associated with lymphocytes. CX3CR1 is also expressed by monocytes and plays an important role in their survival. | |||||||||||||||||||||||||||||||||
| 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 | Tissue homogenates, cell lysates, and other biological fluids |
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4.7 ★★★★★
Based on 13 reviews
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Product Reviews
★★★★★ 5
love shifters
Format: Kindle
Great series with shifters living with and overcoming traumatic experiences with love, community, compassion, kindness and respect.
A great series for these times.
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Reviewed in the United States on March 19, 2025
★★★★★ 3
Heather's story
Format: Paperback
3 stars - This is the second book in the Elena Abbott Shifters series.
In Book Two, we get to know Heather McKenna. She was cursed by a witch, paid by her mother, so she couldn’t shift. Heather was told her whole life that being a shifter was abnormal and made her a monster. She was made to feel ashamed and unworthy. She didn’t find acceptance with her human family or with other wolf shifters when she finally left to forge her own path in the world. Her pack put her at the very bottom of the hierarchy and treated her poorly because what good is a wolf shifter who can’t shift?
When this book starts, we learn that Wren, her new Alpha, has helped her release her wolf from the curse. Heather is having a difficult time accepting herself and finding peace with her wolf. She receives a call from her sister informing her that their mother is in the hospital and she needs to come. Their parting wasn’t on good terms. Heather hopes to find closure with her mom and sister and decides to go, despite Wren’s objections.
V is a member of the Raines pack and also the child of the pack Alpha. Just like in the first book, we discover that this pack is governed by a strict hierarchy of dominance and submission. The Alpha’s word is law. V and their father do not see eye to eye. V’s non-binary identity and attraction to women are not respected by their father, who insists that they mate with Sinclair, the pack enforcer. As one might expect, Sinclair is terrible, and V wants nothing to do with him.
V and Heather meet when V is called to the hospital to handle a potential rogue wolf, who turns out to be Heather. She is struggling to control her emotions and her wolf during the full moon. V immediately recognizes that Heather is their true fated mate. However, it takes Heather, who is not as in tune with her alter ego, a bit longer to come around.
I enjoyed the story, but I must admit that it dragged a bit for me because Heather’s self-esteem is extremely low. She believes she is unworthy of anyone’s love or respect. She thinks she is broken and burdensome. She takes responsibility for and apologizes for everything, most of which she is not responsible for. One of my least favorite tropes is one-sided decision-making, like when a character decides, “I am not worth all the trouble I am causing. They can do better than me. I am doing them a favor by leaving.” Then they leave without giving the other person in the relationship a chance to make any decisions for themselves. Heather’s negative self-talk persists for at least half the book. I found myself skimming through these internal monologues because my tolerance for them is low.
The book picks up in the second half, and I enjoyed the action scenes and the ending. I love how Wren’s new pack is starting to fill up with like-minded wolves, and I look forward to the next installment in the series.
Thank you Netgalley and Bold Strokes Books for the ARC and the opportunity to leave a review.
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Reviewed in the United States on October 16, 2023
★★★★★ 5
Thanks Elena Great Sequel
Format: Kindle
Elena, you did a marvelous job with your writing and character development. It is obvious you put your heart and soul into the story. I look forward to reading more of your work. Please keep it going strong.
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Reviewed in the United States on November 26, 2023
★★★★★ 5
Paranormal Werewolf Shifter Romance
Format: Paperback
Really good second book in this series. This is the story about Heather whose a werewolf shifter who couldn't shift, but gets help from a witch and an alpha werewolf that brings out her wolf , but she goes rogue after shifting and runs. When Heather and V meet they know that they are each others mate, but there's problems from the beginning. There's a lot of violence, drama and angst in this story, so I would say buckle up for a wild ride.. I would definitely recommend this book and I look forward to what's next in this series from this author.
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Reviewed in the United States on October 27, 2023
★★★★★ 4
4 Stars
Format: Paperback
This is a shifter romance that features a non-binary wolf whose father, an Alpha, is stuck in the old ways of running a pack. In comes Heather, featured within the previous installment of the series, who doubts herself and has self-hatred due to her strained relationship with her mother. There are werewolves, witches, dominants and submissives. It's a slow building romance at first and then full steam ahead. For those who are triggered by self-doubt, you may want to pass this up as well as those who hate the mention of the phrase mate or my mate. It is a good second installment of the series with enough open ended content to have a third book on the horizon.
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Reviewed in the United States on December 22, 2023