LBIS™ Mouse IL-12 ELISA Kit
- for Immunochemistry
- Manufacturer :
- FUJIFILM Wako Pure Chemical Corporation
- Storage Condition :
- Keep at 2-10 degrees C.
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96Tests
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In stock in Japan |
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Kit component
96 tests
| Antibody-coated Plate | 96 wells/1 plate |
|---|---|
| Mouse IL-12 Standard | 1 bottle |
| Buffer | 60 mL/1 bottle |
| Biotin-conjugated Antibody | 1 bottle |
| Peroxidase-conjugated Streptavidin Solution | 100 μL/1 bottle |
| TMB Solution | 12 mL/1 bottle |
| Stop Solution | 12 mL/1 bottle |
| Wash Solution (10x) | 100 mL/1 bottle |
| Plate Seal | 4 sheets |
Product Overview
Interleukin-12 (IL-12) is a heterodimeric cytokine composed of two subunits with molecular weights of 35,000 (p35) and 40,000 (p40). It is also known as natural killer cell stimulatory factor (NKSF) and cytotoxic lymphocyte maturation factor (CLMF). IL-12 is produced by macrophages, B cells, and dendritic cells. It plays an important role in inducing cell-mediated immune responses by promoting interferon-γ (IFN-γ) production in T cells and innate lymphoid cells.
LBIS™ Mouse IL-12 ELISA Kit is a sandwich ELISA kit for the quantitative measurement of mouse IL-12. This kit enables highly sensitive quantification of IL-12 in biological samples. The standard included in the kit is a recombinant protein produced in Escherichia coli.
Kit Performance
| Assay target | IL-12 (IL-12p70) *1 |
|---|---|
| Calibration curve range | 2.87 - 700 pg/mL |
| Analysis sample | Mouse Serum/Plasma (EDTA) /Cell culture supernatant*2 |
| Sample amount | 100 μL |
| Measurement duration | Approx 4 hours and 50 minutes |
| Wavelength | Primary wavelength : 450 nm Reference wavelength : 620 nm |
When using cell culture supernatant as a sample, please make sure to perform recovery test and dilution linearity test on the culture medium before use.
Example of Calibration Curve
Data
Repeatability (within run precision)
Repeatability was conducted by measuring fivefold assays of mouse serum samples at two concentrations.
| n＼ID | ID1 (pg/mL) | ID2 (pg/mL) |
|---|---|---|
| 1 | 99.0 | 237 |
| 2 | 94.0 | 218 |
| 3 | 91.8 | 214 |
| 4 | 95.8 | 221 |
| 5 | 96.0 | 229 |
| mean | 95.3 | 224 |
| SD | 2.7 | 9.2 |
| CV(%) | 2.8 | 4.1 |
- [Result]
- The CV (%) of mouse serum was 2.8-4.1%, indicating good repeatability.
Reproducibility (between run precision)
Reproducibility was conducted by measuring human serum samples at three concentrations in triplicate each for four days.
| Day＼ID | ID3 (pg/mL) | ID4 (pg/mL) | ID5 (pg/mL) |
|---|---|---|---|
| 1 | 54.4 | 164.00 | 242 |
| 2 | 61.4 | 173 | 257 |
| 3 | 55.6 | 169 | 258 |
| 4 | 63.4 | 178 | 254 |
| mean | 58.7 | 171 | 253 |
| SD | 4.4 | 5.9 | 7.4 |
| CV(%) | 7.5 | 3.5 | 2.9 |
- [Result]
- The CV (%) of mouse serum was 2.9-7.5%, indicating good reproducibility.
Dilution Linearity Test
Mouse serum and plasma (EDTA), culture medium, as well as cell culture supernatant spiked with the standard, were serially diluted twofold using the provided buffer and measured in duplicate to evaluate linearity.
Mouse serum/plasma (EDTA)
Culture medium (0%/10% FBS)
Raw264.7 supernatant + LPS (Lipopolysaccharide) stimulation
- [Result]
- All measurement results showed good linearity within the measurement range.
Spike and Recovery Test
Standard solutions at three or four concentrations were added to mouse serum/plasma (EDTA), as well as culture medium, then spike recovery tests were conducted, with each measurement performed in duplicate.
Mouse serum
| ID | Spiked amount (pg/mL) |
Measurement value (pg/mL) |
Recovery amount (pg/mL) |
Recovery (%) |
|---|---|---|---|---|
| 1 | - | 38.2 | - | - |
| 50.0 | 82.7 | 44.5 | 89.0 | |
| 150 | 192 | 154 | 103 | |
| 350 | 404 | 366 | 105 | |
| 2 | - | 10.7 | - | - |
| 20.0 | 31.9 | 21.2 | 106 | |
| 60.0 | 71.7 | 61.0 | 102 | |
| 140 | 148 | 137 | 97.9 | |
| mean | 100.3 | |||
Mouse plasma (EDTA)
| ID | Spiked amount (pg/mL) |
Measurement value (pg/mL) |
Recovery amount (pg/mL) |
Recovery (%) |
|---|---|---|---|---|
| 1 | - | 41.6 | - | - |
| 50.0 | 88.4 | 46.8 | 93.5 | |
| 150 | 202 | 160 | 107 | |
| 350 | 417 | 375 | 107 | |
| 2 | - | 12.4 | - | - |
| 20.0 | 32.7 | 20.3 | 102 | |
| 60.0 | 69.0 | 56.6 | 94.4 | |
| 140 | 151 | 139 | 99.3 | |
| mean | 100.4 | |||
- [Result]
- Good recovery rates were confirmed.
Culture medium without FBS
| Spiked amount (pg/mL) |
Measurement value (pg/mL) |
Recovery amount (pg/mL) |
Recovery (%) |
|---|---|---|---|
| 35 | 28.61 | 28.6 | 81.8 |
| 70 | 56.72 | 56.7 | 81.0 |
| 140 | 115.19 | 115.2 | 82.3 |
| 280 | 219.58 | 219.6 | 78.4 |
| mean | 80.9 | ||
- [Result]
- In all cases, the mean recovery was 80% or higher.
Correlation with conventional product
For serum samples of inflammatory mouse model, measurements were conducted using this kit and competitor product A. The measurement values obtained using both products were then compared.
Sample measurement value
Correlation
- [Result]
- Good correlation was confirmed.
Detection Performance Comparison
Mouse serum samples were measured and compared using this kit and the competitor.
| Fujifilm Wako | Company A | |||||||||
|---|---|---|---|---|---|---|---|---|---|---|
| ID＼n | Measurement value (pg/mL) |
mean | SD | CV (%) |
Measurement value (pg/mL) |
mean | SD | CV (%) |
||
| 1 | 2 | 1 | 2 | |||||||
| 1 | 10.9 | 10.2 | 10.6 | 0.51 | 4.8 | N.D. | N.D. | - | - | - |
| 2 | 11.1 | 10.9 | 11.0 | 0.08 | 0.76 | N.D. | N.D. | - | - | - |
| 3 | 10.1 | 9.63 | 9.87 | 0.34 | 3.4 | N.D. | N.D. | - | - | - |
| 4 | 9.75 | 8.56 | 9.15 | 0.84 | 9.2 | N.D. | N.D. | - | - | - |
| 5 | 9.15 | 8.44 | 8.80 | 0.50 | 5.7 | N.D. | N.D. | - | - | - |
N.D.: Below the lower limit of quantification
- [Result]
- This product was able to detect IL-12 even in samples that were below the lower limit of quantification of another manufacturer's kit.
FAQ
About samples
- What anticoagulants should I use?
- We recommend EDTA or as anticoagulant.
- How should I store and prepare the samples?
- Samples should be immediately assayed or stored below -35°C until assay. Before starting assay, shake thawed samples sufficiently. Do not repeat freeze-and-thaw cycles. Dilution of a sample should be made in a PP or PE test tube using the provided buffer prior to adding them to wells.
About kit usage
- What instruments, and equipment are required for the assay using this kit?
- The instruments and equipment required for the use of this kit are listed below;
- Deionized water (or distilled water)
- Test tubes for preparation of standard solution series
- Glassware for dilution of Wash Solution (10x) (a graduated cylinder, a bottle)
- Pipettes (disposable tip type). One should be able to deliver 50 µL precisely, and another for 100 – 1,000 µL
- Syringe-type repeating dispenser like Eppendorf multipette plus which can dispense 100 µL
- Paper towel to remove washing buffer remaining in wellsv
- A vortex-type mixer
- A shaker for 96 well-plate (600 – 1,200 rpm)
- An automatic washer for 96 well-plate (if available), or a wash bottle with a jet nozzle
- A 96 well-plate reader (450 ± 10 nm, 620 nm :600 - 650 nm)
- Software for data analysis
- How should I use the mouse IL-12 standard?
- The amount of purified water to be added varies by lot. Please reconstitute the mouse IL-12 standard with the amount of purified water specified in the “Reconstitution of standard” section on this product page to prepare the original standard solution (7 ng/mL). Then, dilute the solution using the kit-provided buffer that has been equilibrated to room temperature.
- What are the storage conditions for each reagent?
- Buffer & TMB Solution
Use only volume you need for your assay. Remaining reagents should be stored at 2-10°C closing the cap tightly. Once opened, we recommend using as soon as possible to avoid influence by environmental condition.
Stop Solution
Close the cap tightly and store at 2-10°C.
Mouse IL-12 Standard & Biotin-conjugated Antibody
The reconstituted standard solution (original standard solution) should be used within 2 weeks if stored in a refrigerator. Dispose the remaining diluted standard solutions after use.
Peroxidase-conjugated Streptavidin Solution
Remaining working solution (already diluted) should be disposed. The rest of the undiluted solution (unused): close the cap tightly and store at 2-10°C. Once opened, we recommend using as soon as possible to avoid influence by environmental condition.
Prepare reagent solutions in appropriate volume for your assay.
Overview / Applications
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Manufacturer Information
Alias
- 638-40841 (Previous code)
AKMIL12-011 (Previous code)
For research use or further manufacturing use only. Not for use in diagnostic procedures.
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