LBIS™ Mouse/Rat High Molecular Weight Adiponectin 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
For 96 tests
| (A) Antibody-coated Plate | 96 wells (8×12) / 1 plate |
|---|---|
| (B) Adiponectin Standard | 200 μL / 1 bottle |
| (C) Buffer | 60 mL / 1 bottle |
| (D) Peroxidase-conjugated Antibody Solution | 100 μL / 1 bottle |
| (F) TMB Solution | 12 mL / 1 bottle |
| (H) Stop Solution | 12 mL / 1 bottle |
| (I) Wash Solution (10×) | 100 mL / 1 bottle |
| Plate Seal | 3 sheets |
Product Overview
Adiponectin is a cytokine secreted by adipocytes and plays a role in regulating lipid metabolism and insulin sensitivity and is known for its antidiabetic, anti-atherosclerotic, and anti-inflammatory effects. In the bloodstream, adiponectin exists in multimeric forms: trimers (low molecular weight, LMW), hexamers (medium molecular weight, MMW), and higher-order complexes of 12 to 18 monomers (high molecular weight, HMW). Circulating HMW adiponectin levels, rather than total adiponectin, are considered a more accurate indicator of physiological parameters such as BMI, sex, weight loss, glucose tolerance, hepatic insulin sensitivity, and the presence of metabolic syndrome or type 2 diabetes.
This kit is a sandwich ELISA kit designed for the quantitative determination of HMW adiponectin in mouse and rat samples. All reagents are provided in ready-to-use solution form.
Kit Performance
| Analysis sample | Mouse Serum/Plasma Rat Serum/Plasma Culture medium |
|---|---|
| Calibration curve range | 3.13-200 ng/mL |
| Sample volume | 10 μL (diluted 25-50 fold) |
| Measurement duration | Approx. 4 hours |
| Detection method | Colorimetric (Primary : 450 nm / Refference : 620 nm) |
Example of Calibration Curve
Data
Repeatability (within run precision)
Repeatability was conducted by measuring five fold assays of samples at two concentrations with this product.
| n＼ID | A (ng/mL) | B (ng/mL) |
|---|---|---|
| 1 | 29.5 | 129 |
| 2 | 30.7 | 125 |
| 3 | 29.8 | 128 |
| 4 | 29.0 | 126 |
| 5 | 29.6 | 126 |
| mean | 29.7 | 127 |
| SD | 0.622 | 1.64 |
| CV(%) | 2.1 | 1.3 |
[Result]
The CV (%) of samples was 1.3-2.1%, indicating good repeatability.
Reproducibility (between run precision)
Reproducibility was conducted by measuring samples at three concentrations each for four days with this product.
| Day＼ID | C (ng/mL) | D (ng/mL) | E (ng/mL) |
|---|---|---|---|
| 1 | 196 | 126 | 62.5 |
| 2 | 192 | 130 | 59.1 |
| 3 | 196 | 125 | 60.7 |
| 4 | 190 | 125 | 60.3 |
| mean | 194 | 127 | 60.7 |
| SD | 3.00 | 2.38 | 1.41 |
| CV(%) | 1.6 | 1.9 | 2.3 |
[Result]
The CV (%) of samples was 1.6-2.3%, indicating good reproducibility.
Dilution Linearity Test
Samples spiked with the standard, were serially diluted twofold with buffer in this kit and measured in duplicate to assess linearity.
[Result]
All measurement results showed good linearity within the measurement range.
Spike and Recovery Test
Standard solutions at three concentrations were added to samples. Next, spike recovery tests were conducted, with each measurement performed in duplicate.
| Spiked amount (ng/mL) |
Measurement value (ng/mL) |
Recovery volume (ng/mL) |
Recovery rate (%) |
|
|---|---|---|---|---|
| Sample H | 0.0 | 68.5 | - | - |
| 35.0 | 103 | 34.5 | 98.6 | |
| 65.0 | 132 | 63.5 | 97.7 | |
| 95.0 | 165 | 96.9 | 102 | |
| Average | 99.4 | |||
| Sample I | 0.0 | 23.3 | - | - |
| 18.0 | 40.3 | 17.0 | 94.4 | |
| 26.0 | 50.6 | 27.3 | 105 | |
| 32.0 | 55.5 | 32.2 | 101 | |
| Average | 100 | |||
[Result]
Good recovery rates were confirmed.
Cross-Reactivity with Similar Proteins
Using this product, cross-reactivity was assessed by measuring structurally related proteins at each concentrations.
| Species | Similar Proteins | Concentrations (ng/mL) |
Measurement value (ng/mL) |
Cross-Reactivity (％) |
|---|---|---|---|---|
| Mouse | Adiponectin (HMW) | 1,000 | 1,000 | 100 |
| Adiponectin (Hexamer) | 1,000 | <50 | <5 | |
| Adiponectin (Trimer) | 1,000 | N.D. | - | |
| Adiponectin (Monomer) | 1,000 | N.D. | - | |
| MCH | 1,000 | N.D. | - | |
| TNF-α | 1,000 | N.D. | - | |
| INF-γ | 1,000 | N.D. | - | |
| Insulin | 1,000 | N.D. | - | |
| Leptin | 1,000 | N.D. | - | |
| Rat | Adiponectin (HMW) | 1,000 | 1,000 | 100 |
| Adiponectin (Monomer) | 1,000 | N.D. | - | |
| TNF-α | 1,000 | N.D. | - | |
| INF-γ | 1,000 | N.D. | - | |
| Insulin | 1,000 | N.D. | - |
Measurement Using Actual Samples
Adiponectin in serum from mouse/rat
Examples of serum adiponectin measurements in non-fasted mice and rats are shown below.
| Species | Strains | Age (week) |
Sex | Number of samples |
Measurement value (ng/mL) |
SD |
|---|---|---|---|---|---|---|
| Mouse | BALB/c | 6 | Male | 10 | 2,369 | 743 |
| ICR | 6 | Male | 10 | 2,119 | 802 | |
| BKS.Cg-+Leprdb/+Leprdb/ | 12 | Male | 10 | 1,802 | 875 | |
| KKAy/Ta | 12 | Male | 16 | 2,141 | 937 | |
| B6.V-Kepob | 12 | Male | 10 | 1,229 | 437 | |
| Rat | CD (SD) | 8 | Male | 9 | 3,220 | 670 |
| GK | 12 | Male | 10 | 3,512 | 304 | |
| ZUC-Leprfa | 8 | Male | 10 | 1,991 | 1,120 |
- Sample:
- Serum
- Blood Collection Conditions:
- Fasting: None (non-fasted)
Anesthesia: Isoflurane
Sampling site: Heart (abdominal aorta for KKAy/Ta mice) - Housing Conditions:
- KKAy/Ta mice: Housed individually
HMW Adiponectin in male and female mice
Examples of serum HMW adiponectin measurements in male and female mice are shown below.
- Sample:
- Serum
- Blood Collection Conditions:
- Fasting: 24 hours
Anesthesia: Isoflurane
Sampling site: Heart - Food
- Normal Food (CRF-1)
- Housing Conditions:
- Group-Housing
Changes in HMW Adiponectin Following Pioglitazone Administration
Pioglitazone is known to improve insulin resistance by increasing adiponectin expression. The figure shows HMW adiponectin levels in mice before and after pioglitazone administration, as well as in saline-treated control mice.
< Drug administration >
- Species:
- Mouse (KKAy/Ta, Male, 7-weeks old)
- Food:
- CE-2 supplemented with 20% soybean oil
- Before administration:
- (n=12)
- Control:
- 10 mL/kg Saline (n=6)
- Test:
- 10 mg/10 mL/kg Pioglitazone (n=6)
FAQ
About samples
- What should I do with handling blood sample?
To avoid influence of blood (high lipid or hemolysis, etc.), if your original samples have heavy chyle or hemolysis, do not use them for assay.
Samples should be immediately assayed or stored below -35°C for several days. Defrosted samples should be mixed thoroughly for best results. If you have to store assay samples for a longer period, snap-freeze samples and keep them below -35°C. Avoid repeated freezing and thawing.
About kit usage
- What instruments, and equipment are required for the assay using this kit?
- Distilled water or deionized water
- Test tubes for preparation of standard solution series.
- Glassware for dilution of Wash Solution (10x) (a graduated cylinder and a bottle)
- Pipettes (disposable tip type) One should be able to deliver 5-10 µL precisely, and another for 100 - 500 µL.
- Syringe-type repeating dispenser like Eppendorf multi-pipette plus which can dispense 50 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 washing bottle with a jet nozzle.
- A 96 well-plate reader (450 nm ± 10 nm, 620 nm (600 - 650 nm)).
- Software for data analysis.
- What are the precautions for the solution preparation or measurement?
Please read the instruction manual carefully before measurement to confirm sample conditions, measurement conditions, and measurement methods.
Allow the plate and reagents to equilibrate to room temperature (20°C - 25°C) for approximately 2 hours.
Dilute the wash solution 10-fold with purified water that has been brought to room temperature.
Dilute the standard solution with buffer in this kit that has been brought to room temperature.
- What are the storage conditions guidelines for each reagent?
(A) Antibody-coated Plate
If seal is not removed, put the strip back in a plastic bag with zip-seal originally used for well-plate container and store at 2°C - 10°C.
(B) Adiponectin Standard
Do not allow the undiluted solution to return to room temperature. Immediately fasten the cap tightly and store at 2°C - 10°C. Standard solutions prepared above should be used as soon as possible, and should not be stored. Dispose remaining prepared solution.
(C) Buffer & (F) TMB Solution
Use only volume you need for your assay. Remaining reagents should be stored at 2°C - 10°C fastening the cap tightly. Once opened, we recommend using as soon as possible to avoid influence by environmental condition.
(D) Peroxidase-conjugated Antibody Solution
Unused working solution (already diluted) should be disposed. Undiluted solution should be fastened the cap and stored at 2°C - 10°C.
(H) Stop Solution
Remaining solution should be fastened the cap and stored at 2°C - 10°C.
(I) Wash Solution (10x)
Undiluted solution should be fastened the cap and stored at 2°C - 10°C. Dispose any unused diluted wash solution.
Overview / Applications
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Manufacturer Information
Alias
- 634-13071(Previous Code)
AKMAN-011(Previous Code)
For research use or further manufacturing use only. Not for use in diagnostic procedures.
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