| Field | Specification |
|---|---|
| Target | |
| Alternative names | Growth/differentiation factor 8; GDF-8; Myostatin; Mstn; Gdf8 |
| Gene ID | |
| UniProt # | |
| Reactivity | |
| Applications | |
| Immunogen | Expression system for standard: NS0; Immunogen sequence: D268-S376 |
| Expression system | |
| Expression region | |
| Assay type | |
| Sample type(s) | Cell Culture Supernatant, Serum, Plasma |
| Sensitivity | |
| Detection range | |
| Assay time | |
| Storage | |
| Catalog no. (Mfr.) | |
| Main SKU |
Assay Principle
This sandwich ELISA quantifies mouse growth/differentiation factor 8 (gene Mstn) in cell culture supernatants, serum and plasma (heparin or EDTA). Strips are pre-coated with a capture antibody. After the sample is added, a biotinylated detection antibody binds the captured protein, and an avidin–biotin–peroxidase complex supplies the enzyme. TMB develops color in proportion to the bound target; the reaction is stopped and read at 450 nm, and concentrations are read from the standard curve.
Myostatin (MSTN, also called GDF8) is a TGF-β family growth factor made mainly by skeletal muscle. It acts as a negative regulator of muscle growth, and loss of myostatin causes marked muscle enlargement in mice, cattle and other animals, making it a major target of research on muscle wasting and muscle mass.
Kit Components
| Anti-Mouse MSTN Pre-coated 96-well strip microplate | 1 plate (96 wells) |
|---|---|
| Mouse MSTN Standard | 2 vials, 10 ng/tube |
| Mouse MSTN Biotinylated antibody (100x) | 100 µL |
| Avidin-Biotin-Peroxidase Complex (100x) | 100 µL |
| Sample Diluent | 30 mL |
| Antibody Diluent | 12 mL |
| Avidin-Biotin-Peroxidase Diluent | 12 mL |
| Color Developing Reagent (TMB) | 10 mL |
| Stop Solution | 10 mL |
| Wash Buffer (25x) | 20 mL |
| Adhesive plate sealers | 4 |
Also required (not supplied): microplate reader for 450 nm; incubator; automated plate washer (optional); precision pipettes for 0.5 µL through 1 mL volumes of aqueous solutions; multichannel pipettes (recommended for many samples); deionized or distilled water; 500 mL graduated cylinders; test tubes for dilution.
Kit components are not sold separately.
Performance Data
| Species | Mouse |
|---|---|
| Sample types | Cell culture supernatants, serum and plasma (heparin or EDTA) |
| Assay range | 31.2 pg/mL–2,000 pg/mL |
| Sensitivity | <10 pg/mL |
| Standard | Recombinant mouse Mstn (Asp268–Ser376), expressed in NS0 |
| Cross-reactivity | None detected against related proteins in supplier testing |
| Intra-assay CV | 4.0–5.6% (3 samples, n = 16 each) |
| Inter-assay CV | 5.2–6.8% (3 samples, n = 24 each) |
| Format | 96 wells, removable strips |
Sample dilution: start at 1:50. In the supplier’s tests, serum and plasma samples measured about 20 ng/mL.
Example standard curve (OD450, pg/mL): 0 = 0.025; 31.2 = 0.057; 62.5 = 0.095; 125 = 0.158; 250 = 0.303; 500 = 0.599; 1000 = 1.098; 2000 = 2.063. Run a new standard curve on every plate.
Storage & Handling
Store at 4 °C for up to 6 months or −20 °C for up to 12 months. Avoid repeated freeze–thaw cycles. Ships on gel ice and tolerates up to 3 days at room temperature; freeze on arrival.
Safety
Sample data
| Concentration (pg/ml) | 0 | 31.2 | 62.5 | 125 | 250 | 500 | 1000 | 2000 |
| O.D. | 0.025 | 0.057 | 0.095 | 0.158 | 0.303 | 0.599 | 1.098 | 2.063 |
Intra/inter assay consistency
| Intra-Assay Precision | Inter-Assay Precision | |||||
|---|---|---|---|---|---|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 16 | 16 | 16 | 24 | 24 | 24 |
| Mean (pg/ml) | 51 | 227 | 971 | 59 | 238 | 984 |
| Standard deviation | 2.04 | 11.12 | 54.38 | 4.01 | 12.38 | 58.06 |
| CV (%) | 4.0% | 4.9% | 5.6% | 6.8% | 5.2% | 5.9% |
Kit components
Description|Quantity Pre-coated 96-well strip microplate|1 Standard|2 vials Biotinylated antibody (100x)|100ul Avidin-Biotin-Peroxidase Complex (100x)|100ul Sample Diluent|30ml Antibody Diluent|12ml Avidin-Biotin-Peroxidase Diluent|12ml Color Developing Reagent (TMB)|10ml Stop Solution|10ml Wash Buffer (25x)|20ml Adhesive plate sealers|4Materials required but not provided
- Microplate Reader capable of reading absorbance at 450nm.
- Incubator.
- Automated plate washer (optional).
- Pipettes and pipette tips capable of precisely dispensing 0.5 µl through 1 ml volumes of aqueous solutions.
- Multichannel pipettes are recommended for large amount of samples.
- Deionized or distilled water.
- 500ml graduated cylinders.
- Test tubes for dilution.
Activating reagent preparation
Myostatin/GDF8 is mostly contained as inactive form in samples, please activate it before assay. Don't activate recombinant Myostatin/GDF8.
Solution A: 1N HCI: add 8.33ml of 12N HCI into 91.67ml of H2O.
Solution B: 1.2N NaOH/0.5M HEPES: add 12ml of 10N NaOH and 11.9g HEPES into 75ml of H2O, add H2O to adjust volume to 100ml.
Sample activation procedure
Activate the sample
Cell culture supernatants: add activating reagent pro rata, i.e. add 50μl of Solution A into 100μl of sample, 10 min later, add 50μl of Solution B. PH 7.0-7.6.
Serum, plasma(heparin, EDTA): add activating reagent pro rata, i.e. add 20μl of Solution A into 40μl of sample, 10 min later, add 20μl of Solution B. PH 7.0-7.6.
It is unnecessary to activate the recombinant Myostatin/GDF8.
Sample was diluted partly after adding activating reagent, so please pay attention to this when calculate target protein concentration.
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