LBIS (TM) Mouse IL-6 ELISA Kit
- for Immunochemistry
- Manufacturer :
- FUJIFILM Wako Pure Chemical Corporation
- Storage Condition :
- Keep at 2-10 degrees C.
- GHS :
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96Tests
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In stock in Japan |
※Check availability in the US with the distributor.
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Kit component
96 tests
| Antibody-coated Plate | 1 plate |
|---|---|
| Mouse IL-6 Standard (Freeze-dried) | 2 vials |
| Buffer | 30 mL |
| Biotin-conjugated Antibody Solution | 100 μL |
| Peroxidase-conjugated Streptavidin Solution | 100 μL |
| TMB Solution | 12 mL |
| Buffer for Standard and Sample | 30 mL |
| Stop Solution | 12 mL |
| Wash Solution (10x) | 100 mL |
| Plate seal | 4 sheets |
Product Overview
IL-6 (Interleukin-6) is a secreted glycoprotein of 187 amino acids (in case of mice) and it was discovered to be a cytokine that differentiates B cells into antibody-producing cells. It is also known to play an important role in various physiological processes such as immune response, inflammatory response, hematopoiesis, and the proliferation and differentiation of nervous system cells.
After being produced by T cells, monocytes, fibroblasts, endothelial cells, keratinocytes, etc., it binds to the IL-6 receptor, which consists of two chains of IL-6Rα and gp130, and signals are transduced. There is also a study that suggested that a possible correlation exists between IL-6 levels and disease activity in rheumatoid arthritis, and it is also attracting attention in the field of autoimmune diseases such as rheumatoid arthritis and inflammatory diseases.
This kit is a sandwich ELISA system for quantitative measurement of mouse IL-6. It can measure IL-6 in mouse serum, plasma and culture supernatant, specifically and with high sensitively.
Features
- Designed for specific and highly sensitive measurement of IL-6 in mouse serum, EDTA plasma, and cell culture supernatant
- Requires a total measurement duration of 3 hours and 50 minutes
- The standard is a recombinant protein produced by E. coli and is not subject to the Cartagena Protocol
- The standard is divided into two vials to allow for multiple measurements
Kit Performance
| Calibration curve range | 2.05~500 pg/mL |
|---|---|
| Assay target | IL-6 (Interleukin-6) |
| Analysis sample | Mouse serum/plasma (EDTA)/cell culture supernatant |
| Sample volume | 25 μL (2-fold dilution) |
| Measurement duration | 3 hours 50 minutes |
| Detection method | Chromogenic (Primary wavelength 450 nm / reference wavelength 620 nm) |
Example of Calibration Curve
Data
Precision of assay (within assay variation)
Two serum samples, each at a different concentration, were measured in quintuplicate using this kit to confirm reproducibility.
| n＼ID | ID1 (pg/mL) | ID2 (pg/mL) |
|---|---|---|
| 1 | 281 | 18.7 |
| 2 | 285 | 19.1 |
| 3 | 281 | 18.7 |
| 4 | 282 | 18.7 |
| 5 | 291 | 19.6 |
| mean | 284 | 19.0 |
| SD | 4.18 | 0.390 |
| CV(%) | 1.47 | 2.06 |
- [Result]
- The CV (%) was 1.47% and 2.06%, indicating good reproducibility.
Reproducibility (between assay variation)
Three serum samples, each at a different concentration, were measured in triplicate over four consecutive days using this kit.
| Day＼ID | ID3 (pg/mL) | ID4 (pg/mL) | ID5 (pg/mL) |
|---|---|---|---|
| 1 | 246 | 84 | 15.1 |
| 2 | 230 | 78.8 | 13.9 |
| 3 | 254 | 86.2 | 15.3 |
| 4 | 244 | 84.9 | 14.4 |
| mean | 244 | 83.5 | 14.7 |
| SD | 9.90 | 3.23 | 0.658 |
| CV(%) | 4.07 | 3.87 | 4.49 |
- [Result]
- The CV (%) ranged from 3.87% to 4.49%, indicating good reproducibility.
Dilution linearity test
Samples of serum, EDTA plasma, D-MEM medium, and 10% FCS/D-MEM medium, each at two different concentrations, were subjected to a 2-fold serial dilution with sample buffer and measured to confirm linearity (measured in duplicates).
- [Result]
- Good linearity was confirmed for all samples within the calibration curve range.
Spiked recovery test
Standard solutions at four concentrations were added to serum, EDTA plasma, D-MEM medium, and 10% FCS/D-MEM medium samples. Next, spike-recovery tests were conducted, with each measurement performed in duplicate.
| Amount spiked (pg/mL) | Measured value (pg/mL) | Recovery volume (pg/mL) | Recovery rate (%) | |
|---|---|---|---|---|
| Serum | - | 4.8 | - | - |
| 22.8 | 26.9 | 22.1 | 96.9 | |
| 46.0 | 49.0 | 44.2 | 96.0 | |
| 96.4 | 94.9 | 90.1 | 93.4 | |
| 197.0 | 193 | 188 | 95.5 | |
| Average | 95.5 | |||
| Plasma (EDTA) | - | 4.4 | - | - |
| 28.1 | 30.6 | 26.2 | 93.0 | |
| 56.0 | 55.0 | 50.5 | 90.2 | |
| 116 | 107 | 102 | 88.4 | |
| 227 | 206 | 202 | 88.8 | |
| Average | 90.1 | |||
| D-MEM | - | 0.0 | - | - |
| 23.0 | 24.1 | 24.1 | 105.0 | |
| 48.3 | 47.7 | 47.7 | 98.7 | |
| 97.5 | 93 | 93.0 | 95.3 | |
| 194.4 | 205 | 205.1 | 105.5 | |
| Average | 101.1 | |||
| 10% FCS /D-MEM | - | 0.0 | - | - |
| 28.1 | 27.6 | 27.6 | 98.1 | |
| 56.0 | 60.2 | 60.2 | 107.5 | |
| 115.6 | 121 | 120.7 | 104.4 | |
| 227.3 | 237 | 237.2 | 104.3 | |
| Average | 103.6 | |||
- [Result]
- The recovery rates ranged from 88.4% to 107.5%, indicating good performance.
Cross-reactivity with similar proteins
Proteins similar to mouse IL-6, including IL-6 of other species and other interleukins, were tested to determine if they could be detected.
| Proteins | Amount spiked (pg/mL) | Measured value (pg/mL) |
|---|---|---|
| Human IL-6 | 500 | 0.034 |
| Rat IL-6 | 500 | 0.214 |
| Mouse IL-4 | 1,000 | 0.01 |
| Mouse IL-7 | 1,000 | - |
| Mouse gp130 | 1,000 | - |
| Mouse IL-6Rα | 1,000 | - |
-: The values are outside the standard curve.
- [Result]
- Very low reactivity with similar proteins was confirmed.
Correlation with conventional product
For mouse serum samples, measurements were conducted using this kit and competitor product A. The measurement values obtained using both products were then compared.
- [Result]
- A good correlation was confirmed.
Detection Performance Comparison
Measurements were conducted on mouse serum samples using this kit and competitor product A to compare their detectability.
| Sample No. | Fujifilm Wako (pg/mL) | Company A (pg/mL) |
|---|---|---|
| 1 | 1.71 | Not detectable |
| 2 | 6.50 | 5.52 |
| 3 | 6.39 | 9.02 |
| 4 | 7.01 | 7.67 |
| 5 | 9.67 | 7.53 |
| Proportion of samples successfully measured |
4/5 (80%) | 1/5 (20%) |
| Samples measured within the calibration range are indicated in red; samples outside the range are noted as reference values. | ||
- [Result]
- This kit demonstrated high detection sensitivity, successfully detecting four out of five samples within the calibration curve range.
FAQ
About sample
- How should samples be stored?
- For long-term storage of samples, please store below –35℃ until assay. Before starting the assay, thoroughly shake thawed samples. Do not repeat freeze-and-thaw cycles.
- Can I measure heparin plasma or citric acid plasma as a sample?
- Please use only EDTA as anticoagulant.
- How should samples with turbidity or insoluble matter be handled?
- Centrifuge the sample to remove turbidity or insoluble matter where necessary before use in the assay.
About the kit
- How do I calibrate the measured values?
- The measured value obtained by this kit can be converted to the unit concentration based on NIBSC/WHO standard mouse IL-6 (Code: 93/730) by multiplying with the conversion factor specified in a separate sheet according to the following calculation formula.
NIBSC/WHO(93/730) unit concentration (U/mL)=Conversion factor x Measured mouse IL-6 value (pg/mL) ＊The conversion factor is different depending on the lot. Please see the specified conversion factor in the annex document.
- Is it possible to purchase the antibodies provided in this kit?
- These antibodies are not sold separately.
- Can I divide plates?
- It is possible. The plate is designed to allow separation into individual sections of 8 wells each. The kit includes a sufficient amount of standard for this purpose. However, please be aware that if the plate is used in separate measurements, this necessitates redrawing the calibration curve each time, which, in turn, reduces the total number of samples that can be measured.
For single use → 40 samples (80 wells), with 16 wells used for standards and blank.
For two uses → 32 samples (64 wells), with 32 wells used for standards and blank.
About kit usage
- What is the amount of purified water to be added to the standard product?
- Because the volume of deionized water (or distilled water) to be added to the freeze-dried standard differs depending on the lot, see the specified volume in a separate sheet. Be sure to allow the solution to stand for 30 minutes, as accurate values may not be obtained.
Overview / Applications
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For research use or further manufacturing use only. Not for use in diagnostic procedures.
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