LBIS (TM) Human IFN-γ 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 |
|---|---|
| Human IFN-γ 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
Interferon-γ (IFN-γ) is an important cytokine involved in both innate and adaptive immunity. Cells known to produce IFN-γ include natural killer (NK) cells and T cells. Secreted IFN-γ induces a wide range of immune responses, including antiviral activity, activation of macrophages, maturation of dendritic cells, and induction of T-cell differentiation. IFN-γ also plays a significant role in antitumor immunity, with reported effects including suppression of metastasis, induction of apoptosis, and inhibition of angiogenesis.
LBIS™ Human IFN-γ ELISA Kit is a sandwich ELISA kit for the quantitative measurement of human IFN-γ. This kit enables highly sensitive quantification of IFN-γ in biological samples. The standard included in the kit is a recombinant protein produced in Escherichia coli.
Kit Performance
| Assay target | IFN-γ |
|---|---|
| Calibration curve range | 0.768-75.0 pg/mL |
| Analysis sample | Human Serum/Plasma (EDTA, heparin) /Cell culture supernatant* |
| Sample amount | 50 μL (2x dilution) |
| Measurement duration | Approx 2 hours and 20 minutes |
| Wavelength | Primary wavelength : 450 nm Reference wavelength : 620 nm |
Example of Calibration Curve
Data
Repeatability (within run precision)
Repeatability was conducted by measuring fivefold assays of human serum samples at two concentrations.
| n＼ID | ID1 (pg/mL) | ID2 (pg/mL) |
|---|---|---|
| 1 | 26.1 | 1.41 |
| 2 | 26.4 | 1.42 |
| 3 | 26.2 | 1.44 |
| 4 | 26.3 | 1.46 |
| 5 | 26.4 | 1.43 |
| mean | 26.3 | 1.43 |
| SD | 0.13 | 0.019 |
| CV(%) | 0.50 | 1.3 |
- [Result]
- The CV (%) of human serum was 0.50-1.3%, 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 | 29.1 | 4.90 | 0.742 |
| 2 | 29.9 | 4.69 | 0.723 |
| 3 | 31.2 | 4.87 | 0.785 |
| 4 | 29.5 | 4.99 | 0.772 |
| mean | 29.9 | 4.86 | 0.756 |
| SD | 0.91 | 0.13 | 0.028 |
| CV(%) | 3.0 | 2.6 | 3.7 |
- [Result]
- The CV (%) of human serum was 2.6-3.7%, indicating good reproducibility.
Dilution Linearity Test
Human serum and plasma (EDTA/heparin), 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.
Human serum/plasma (EDTA, heparin)
Human PBMC supernatants ± PHA stimulation (in RPMI-1640 medium with 10% FBS)
Human PBMC supernatants stimulated with PHA (phytohemagglutinin, simulating IFN-γ secretion) for 3 or 6 days were measured at 1:100 dilution.
- [Result]
- All measurement results showed good linearity within the measurement range.
Spike and Recovery Test
Standard solutions at three concentrations were added to human serum/plasma (EDTA/heparin), then spike recovery tests were conducted, with each measurement performed in duplicate.
Human serum
| Spiked amount (pg/mL) |
Measurement value (pg/mL) |
Recovery amount (pg/mL) |
Recovery (%) |
|---|---|---|---|
| - | 0.516 | - | - |
| 1.00 | 1.60 | 1.08 | 108 |
| 4.00 | 4.50 | 3.98 | 99.5 |
| 16.0 | 16.4 | 15.9 | 99.4 |
| mean | 102 | ||
Human plasma (EDTA)
| Spiked amount (pg/mL) |
Measurement value (pg/mL) |
Recovery amount (pg/mL) |
Recovery (%) |
|---|---|---|---|
| - | 0.708 | - | - |
| 1.00 | 1.76 | 1.05 | 105 |
| 4.00 | 4.47 | 3.76 | 94.0 |
| 16.0 | 15.1 | 14.4 | 90.0 |
| mean | 96.3 | ||
Human plasma (heparin)
| Spiked amount (pg/mL) |
Measurement value (pg/mL) |
Recovery amount (pg/mL) |
Recovery (%) |
|---|---|---|---|
| - | 0.618 | - | - |
| 1.00 | 1.62 | 1.00 | 100 |
| 4.00 | 4.20 | 3.58 | 89.6 |
| 16.0 | 14.5 | 13.9 | 86.8 |
| mean | 92.2 | ||
- [Result]
- Good recovery rates were confirmed.
Cross-reactivity with similar proteins
Proteins similar to human IFN-γ, including IFN-γ of other species and human IFN-γR1 (IFN-γ Receptor 1), were tested to determine if they could be detected.
| Spiked amount (ng/mL) |
Measurement value (ng/mL) |
Cross-reactivity (%) |
|
|---|---|---|---|
| Mouse IFN-γ | 2 | N.D. | - |
| 10 | N.D. | - | |
| 50 | N.D. | - | |
| Rat IFN-γ | 2 | N.D. | - |
| 10 | N.D. | - | |
| 50 | N.D. | - | |
| Dog IFN-γ | 2 | N.D. | - |
| 10 | 0.0152 | 0.15 | |
| 50 | 0.0983 | 0.20 | |
| Bovine IFN-γ | 2 | N.D. | - |
| 10 | N.D. | - | |
| 50 | N.D. | - | |
| Monkey IFN-γ | 2 | 0.0206 | 1.03 |
| 10 | 0.1527 | 1.53 | |
| Rabbit IFN-γ | 2 | N.D. | - |
| 10 | N.D. | - | |
| 50 | N.D. | - | |
| Human IFN-γR1 | 2 | N.D. | - |
| 10 | N.D. | - | |
| 50 | N.D. | - |
N.D.: Below the lower limit of quantification
Red: Values outside the quantification range; shown as reference
- [Result]
- Very low reactivity with similar proteins was confirmed.
Example of Sample Measurement
Human serum and plasma (EDTA/heparin) samples were measured and compared using this kit and the competitor.
Human serum
| ID＼n | Fujifilm Wako (pg/mL) |
Company A (pg/mL) |
|---|---|---|
| 1 | 0.821 | N.D. |
| 2 | 1.21 | N.D. |
| 3 | N.D. | N.D. |
| 4 | N.D. | N.D. |
| 5 | 2.02 | N.D. |
| 6 | N.D. | N.D. |
| 7 | 2.02 | N.D. |
| 8 | 1.43 | N.D. |
| 9 | 0.974 | N.D. |
| 10 | 1.71 | N.D. |
Human plasma (heparin)
| ID＼n | Fujifilm Wako (pg/mL) |
Company A (pg/mL) |
|---|---|---|
| 21 | N.D. | N.D. |
| 22 | N.D. | N.D. |
| 23 | 2.68 | N.D. |
| 24 | 1.28 | N.D. |
| 25 | 1.39 | N.D. |
| 26 | N.D. | N.D. |
| 27 | 0.865 | N.D. |
| 28 | 3.00 | 8.92 |
| 29 | 2.75 | N.D. |
| 30 | 2.44 | N.D. |
N.D.: Below the lower limit of quantification
Human PBMC supernatants ± PHA stimulation (in RPMI-1640 medium with 10% FBS)
| Samples | Fujifilm Wako (pg/mL) | Company A (pg/mL) |
|---|---|---|
| RPMI-1640 + 10% FBS | N.D. | N.D. |
| PHA (-) human PBMC | N.D. | N.D. |
| PHA (+) human PBMC | 12,385 | 16,224 |
N.D.: Below the lower limit of quantification
- [Result]
- This product was able to detect and quantify IFN-γ in human serum/plasma samples in which IFN-γ was not detected using the other manufacturer's product. With both products, IFN-γ levels were below the limit of quantification in the control medium and in culture supernatants of human PBMCs without PHA stimulation. In contrast, elevated IFN-γ levels were confirmed in culture supernatants of PHA-stimulated cells.
FAQ
About samples
- What anticoagulants should I use?
- We recommend using EDTA or heparin as anticoagulant.
- How should I store and prepare the samples?
- Samples should be immediately assayed or stored below -35℃ 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 wells
- 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 human IFN-γ standard?
- The amount of purified water to be added varies by lot. Please reconstitute the human IFN-γ standard with the amount of purified water specified in the “Reconstitution of standard” section on this product page to prepare the original standard solution (750 pg/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℃ 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℃.
Human IFN-γ 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℃. 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
- 631-47891 (Previous Code)
AKH-IFNG (Previous Code)
For research use or further manufacturing use only. Not for use in diagnostic procedures.
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