LabAssay (TM) ATX
- for Cellbiology
- Manufacturer :
- FUJIFILM Wako Pure Chemical Corporation
- Storage Condition :
- Keep at 2-10 degrees C.
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150Tests
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In stock in Japan |
Document
Kit component
Kit Component
| Reacting Solution | 16 mL |
|---|---|
| Substrate | 1 bottle (For 8 mL) |
| Substrate Dissolving Solution | 8 mL |
| ATX Standard | 2 bottles |
| Buffer | 5 mL |
| Stop Solution | 16 mL |
Product Overview
Autotaxin (ATX) is a glycoprotein with a molecular weight of 125 kDa that was isolated as a cell migration promoting factor from the cell culture supernatant of human malignant melanoma cells. It is known that liver damage such as fibrosis causes metabolic inhibition of ATX, which leads to retention of ATX in the blood, resulting in an increase in its blood concentration.
LabAssay™ ATX is a kit used for the determination of ATX in blood (serum and plasma) and cell culture supernatant. With the use of a microplate, this kit provides a convenient method for measuring ATX in samples.
[Note] LabAssay™ series are reagents for research purposes. It cannot be used for diagnostic purposes.
Kit Performance
| Sample | Serum, Plasma (Heparin), Cell culture supernatant* |
|---|---|
| Target animals | Human, Mouse, Rat |
| Calibration curve range | 1.72-55 U/L |
| Sample volume | 10 μL |
| Measurement duration | Approx. 40 min |
| Wavelength | Primary wavelength 546 nm Reference wavelength 700 nm |
* Measurement availability depends on the culture medium.
Example of Calibration Curve
Principle
Lysophosphatidylcholine is hydrolyzed by the lysophospholipase D activity of ATX in the sample to form choline. The resultant choline is oxidized by the action of choline oxidase (COD) to form hydrogen peroxide. The resultant hydrogen peroxide oxidatively condenses N-Ethyl-N-(3-sulfopropyl)-3-methylaniline (TOPS) and 4-aminoantipyrine quantitatively through the action of peroxidase (POD) to produce a blue-violet pigment. The reaction is stopped by addition of the Stop Solution and absorbance is measured to determine ATX activity in the sample.
Data
Precision of assay (within assay variation)
Reproducibility was confirmed by performing quintuplicate assays of serum samples of two concentrations each from humans, mice, and rats with this product.
| n＼ID | Human | Mouse | Rat | |||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Serum 1 | Serum 2 | Plasma 1 | Plasma 2 | Serum 1 | Serum 2 | Plasma 1 | Plasma 2 | Serum 1 | Serum 2 | Plasma 1 | Plasma 2 | |
| 1 | 24.5 | 6.31 | 26.3 | 6.75 | 36.4 | 6.64 | 35.0 | 5.92 | 25.6 | 9.95 | 24.2 | 10.3 |
| 2 | 24.5 | 6.55 | 26.4 | 6.94 | 37.3 | 6.58 | 35.0 | 5.92 | 24.0 | 9.82 | 24.0 | 10.3 |
| 3 | 25.1 | 6.44 | 26.8 | 6.89 | 36.3 | 6.73 | 34.9 | 5.97 | 23.9 | 10.2 | 24.7 | 10.6 |
| 4 | 25.2 | 6.61 | 26.8 | 7.06 | 37.2 | 6.70 | 34.9 | 5.95 | 24.5 | 10.0 | 24.5 | 10.5 |
| 5 | 25.1 | 6.43 | 26.7 | 6.86 | 36.2 | 6.66 | 34.9 | 5.93 | 24.5 | 10.0 | 24.5 | 10.5 |
| mean | 24.9 | 6.47 | 26.6 | 6.90 | 36.7 | 6.66 | 34.9 | 5.94 | 24.5 | 10.0 | 24.4 | 10.4 |
| SD | 0.35 | 0.12 | 0.23 | 0.11 | 0.53 | 0.058 | 0.055 | 0.024 | 0.67 | 0.14 | 0.28 | 0.13 |
| CV(%) | 1.4 | 1.8 | 0.88 | 1.6 | 1.4 | 0.86 | 0.16 | 0.40 | 2.8 | 1.4 | 1.1 | 1.3 |
Unit: U/L
[Result]
The CV (%) of human serum was 0.88-1.8%, mouse serum was 0.16-1.4%, and rat serum was 1.1-2.8%, indicating good reproducibility.
Reproducibility (between assay variation)
Triplicate measurements were performed on human, mouse, and rat serum samples at three concentrations each for four days with this product.
| Day＼ID | Human | Mouse | Rat | |||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Serum 1 | Serum 2 | Serum 3 | Plasma 1 | Plasma 2 | Plasma 3 | Serum 1 | Serum 2 | Serum 3 | Plasma 1 | Plasma 2 | Plasma 3 | Serum 1 | Serum 2 | Serum 3 | Plasma 1 | Plasma 2 | Plasma 3 | |
| 0 | 24.4 | 18.2 | 13.7 | 26.3 | 18.7 | 14.0 | 41.1 | 20.6 | 9.88 | 39.8 | 20.0 | 9.32 | 21.7 | 15.9 | 11.8 | 21.2 | 15.1 | 10.6 |
| 1 | 24.6 | 18.3 | 13.8 | 26.5 | 18.7 | 14.0 | 40.9 | 20.4 | 9.82 | 39.5 | 19.9 | 9.31 | 22.0 | 16.0 | 11.9 | 21.3 | 15.2 | 10.8 |
| 2 | 23.6 | 17.5 | 13.8 | 25.4 | 18.3 | 14.6 | 40.0 | 20.2 | 10.1 | 40.1 | 20.2 | 9.90 | 22.7 | 16.6 | 13.0 | 22.3 | 15.6 | 11.7 |
| 3 | 23.6 | 17.5 | 13.8 | 25.4 | 18.3 | 14.6 | 39.4 | 20.1 | 10.1 | 39.4 | 20.1 | 9.88 | 22.8 | 16.9 | 13.1 | 22.5 | 15.7 | 11.8 |
| mean | 24.1 | 17.9 | 13.8 | 25.9 | 18.5 | 14.3 | 40.4 | 20.3 | 10.0 | 39.7 | 20.1 | 9.60 | 22.3 | 16.4 | 12.5 | 21.8 | 15.4 | 11.2 |
| SD | 0.53 | 0.43 | 0.050 | 0.58 | 0.23 | 0.35 | 0.79 | 0.22 | 0.15 | 0.32 | 0.13 | 0.33 | 0.54 | 0.48 | 0.70 | 0.67 | 0.29 | 0.61 |
| CV(%) | 2.2 | 2.4 | 0.36 | 2.3 | 1.2 | 2.4 | 2.0 | 1.1 | 1.5 | 0.80 | 0.64 | 3.5 | 2.4 | 2.9 | 5.6 | 3.1 | 1.9 | 5.5 |
Unit : U/L
[Result]
The CV (%) of human serum was 0.36-2.4%, mouse serum was 0.64-3.5%, and rat serum was 1.9-5.6%, indicating good reproducibility.
Dilution linearity test
Using saline, human serum/plasma (Heparin), mouse serum/plasma (Heparin), and rat serum/plasma (Heparin) at two different concentrations each, 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 three concentrations were added to human serum/plasma(Heparin), mouse serum/plasma(Heparin) and rat serum/plasma(Heparin). Next, spike-recovery tests were conducted, with each measurement performed in duplicate.
Human
| Amount spiked (U/L) | Measured value (U/L) | Recovery volume (U/L) | Recovery rate (%) | |
|---|---|---|---|---|
| Serum | - | 5.95 | - | - |
| 8.05 | 13.8 | 7.85 | 97.5 | |
| 24.2 | 28.2 | 22.3 | 92.1 | |
| 40.3 | 44.9 | 39.0 | 96.8 | |
| Plasma (Heparin) | - | 7.29 | - | - |
| 7.69 | 14.6 | 7.31 | 95.1 | |
| 23.1 | 28.6 | 21.3 | 92.2 | |
| 38.5 | 46.9 | 39.6 | 103 |
Mouse
| Amount spiked (U/L) | Measured value (U/L) | Recovery volume (U/L) | Recovery rate (%) | |
|---|---|---|---|---|
| Serum | - | 10.1 | - | - |
| 6.08 | 16.8 | 6.70 | 110 | |
| 12.2 | 21.6 | 11.5 | 94.3 | |
| 24.3 | 32.8 | 22.7 | 93.4 | |
| Plasma (Heparin) | - | 11.20 | - | - |
| 6.57 | 17.6 | 6.40 | 97.4 | |
| 13.1 | 23.8 | 12.6 | 95.9 | |
| 26.3 | 36.4 | 25.2 | 95.9 |
Rat
| Amount spiked (U/L) | Measured value (U/L) | Recovery volume (U/L) | Recovery rate (%) | |
|---|---|---|---|---|
| Serum | - | 10.50 | - | - |
| 7.42 | 18.0 | 7.50 | 101 | |
| 14.8 | 24.3 | 13.8 | 93.0 | |
| 29.7 | 38.5 | 28.0 | 94.4 | |
| Plasma (Heparin) | - | 10.7 | - | - |
| 7.48 | 18.0 | 7.30 | 97.6 | |
| 15.0 | 24.2 | 13.5 | 90.2 | |
| 29.9 | 38.1 | 27.4 | 91.6 |
[Result]
Good recovery rate was confirmed.
Correlation with conventional product
Measurements were conducted on the same samples of human serum and plasma (Heparin) using this kit and company R’s product. The measurement values obtained using both products were then compared.
| Sample No. | Serum | Sample No. | Plasma (Heparin) | ||
|---|---|---|---|---|---|
| Fujifilm Wako (U/L) | Company R (ng/mL) | Fujifilm Wako (U/L) | Company R (ng/mL) | ||
| 1 | 8.03 | 209 | 15 | 8.49 | 193 |
| 2 | 8.92 | 202 | 16 | 8.76 | 198 |
| 3 | 7.10 | 162 | 17 | 7.11 | 156 |
| 4 | 7.52 | 148 | 18 | 7.13 | 142 |
| 5 | 6.53 | 133 | 19 | 6.79 | 139 |
| 6 | 9.29 | 201 | 20 | 9.01 | 200 |
| 7 | 7.88 | 184 | 21 | 7.60 | 181 |
| 8 | 7.19 | 176 | 22 | 6.84 | 167 |
| 9 | 6.44 | 174 | 23 | 6.64 | 168 |
| 10 | 11.8 | 281 | 24 | 12.1 | 309 |
| 11 | 6.65 | 140 | 25 | 6.81 | 129 |
| 12 | 6.65 | 144 | 26 | 6.59 | 137 |
| 13 | 7.31 | 166 | 27 | 7.17 | 162 |
| 14 | 7.50 | 145 | 28 | 7.30 | 145 |
Samples measured within the calibration range are indicated in red; samples outside the range were diluted 20-fold and assayed.
[Result]
A good correlation was confirmed. In addition, the other company R’s product required dilution of all samples because the values were outside the calibration curve range. On the other hand, our product could measure within the calibration curve range without diluting the samples.
FAQ
About sample
- What specimens should I use?
- Analyze specimens immediately after collection. If it is difficult to analyze immediately, store frozen until analysis. Hemolysis may influence the assay.
- Is it possible to measure in cell culture supernatant?
- Yes. However, we confirmed that the recovery rate may vary depending on the culture medium in the spiked recovery test, and the high concentration portion may show a high value in the dilution linearity test. If you use cell culture supernatant as a sample, perform a spiked recovery test or dilution linearity test on the medium to be used in advance.
- Which anticoagulants can I use?
- The anticoagulant heparin does not significantly influence the assay when used in normal amounts. Do not use EDTA because it influences the assay.
- What should I do with samples that exceed the measurable range?
- Dilute specimen with saline and repeat the assay if the measured value exceeds the measurable range, and multiply the result by the dilution factor.
- Does ascorbic acid affect the measured values?
- Ascorbic acid does not affect the assay up to 10 mg/dL.
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.
- 96-well microplate (transparent type)
- Micropipette
- Microtube
- Pipette
- Incubator maintained at 37℃
- Plate mixer
- Microplate reader (with 546 nm/700 nm wavelength filter)
- What is the amount of purified water to be added to the standard product?
- Find and check “Reconstitution of standard” on this product page. As the amount of purified water to be added varies by lot, be sure to check it for every lot.
Overview / Applications
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Manufacturer Information
Alias
- autotaxin
For research use or further manufacturing use only. Not for use in diagnostic procedures.
Product content may differ from the actual image due to minor specification changes etc.
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