SKU: 81989900426

Mouse CXCL12 ELISA Kit

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Description

Mouse CXCL12 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 handling and requirements: Plasma: Collect specimens using EDTA or heparin as an anticoagulant. Centrifuge at 1000g for 15 minutes at 2 8C within 30 minutes of collection. Remove the supernatant for analysis or store at 20C

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 handling and requirements:
Plasma: Collect specimens using EDTA or heparin as an anticoagulant.
Centrifuge at 1000×g for 15 minutes at 2-8°C within 30 minutes of collection.
Remove the supernatant for analysis or store at -20°C or -80°C.
Avoid repeated freezing and thawing.

Cell culture supernatant: Centrifuge at 1000×g for 20 minutes.
Remove the supernatant for analysis or store at -20°C or -80°C.
Avoid repeated freezing and thawing.

Pre-Assay Preparation:
1. Remove the test kit from the refrigerator 10 minutes in advance and equilibrate to room temperature.
2. Prepare the gradient working solution of the standard: Add 1 mL of universal diluent to the lyophilized standard, let it stand for 15 minutes to completely dissolve, then gently mix (concentration is 8000 pg/mL).
Then dilute to the following concentrations: 8000 pg/mL, 4000 pg/mL, 2000 pg/mL, 1000 pg/mL, 500 pg/mL, 250 pg/mL, 125 pg/mL, and 0 pg/mL.

Serial Dilution Method: Add 500 μL of universal diluent to each of seven EP tubes.
Pipette 500 μL of the 8000 pg/mL standard working solution into the first EP tube and mix thoroughly to make a 4000 pg/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.

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.
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 a CXC motif chemokine 12 (CXCL12) 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 CXC motif chemokine 12 (CXCL12) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration.
Source Mouse
Synonym Mouse C-X-C motif chemokine 12 ELISA Kit
Detection Type Double antibody sandwich method
Composition
Name 9 6 T  match   set remark
Pre-coating 96 Well plate 8 Hole ×12 Strip without
Standard 2 branch
Dilute as per instructions
Universal diluent
2×20mL
without
concentrate HRP Anti-detection 100×
120uL
Dilute as per instructions
20× Washing liquid
2×10mL
Dilute as per instructions
Bottom thing ( TMB )
10mL
without
Stop liquid
6mL
without
Sealing film
4 Zhang
without
Instructions
1 Share
without
Background CXC chemokine ligand 12 (CXCL12), also known as stromal cell-derived factor 1 (SDF1), is a chemokine protein encoded by the CXCL12 gene. It is ubiquitously expressed in many tissues and cell types. Stromal cell-derived factors 1-α and 1-β are small cytokines belonging to the chemokine family. Members of these chemokines activate white blood cells, typically in response to proinflammatory stimuli such as lipopolysaccharide, TNF, or IL-1. Chemokines are characterized by the presence of four conserved cysteines forming two disulfide bonds. They can be divided into two subfamilies. In the CC subfamily, the cysteine residues are adjacent to each other. In the CXC subfamily, they are separated by an intermediate amino acid. The SDF1 protein belongs to the latter group. Its signaling has been observed in several cancers. Its gene also contains one of 27 SNPs associated with an increased risk of coronary artery disease.
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.
Storage Temp. If the unopened kit is stored at 4°C, the shelf life is 6 months.
Test Range 125-8000pg/mL
Applications Plasma, cell culture supernatant
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Exchange/Return Notes
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SKU: 81989900426

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