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| ID | Components | Size |
| RSP010-C01 | Pre-coated Anti-IL-4 Antibody Microplate | 1 plate |
| RSP010-C02 | Positive Stimulus | 150 μg |
| RSP010-C03 | Biotin-Anti-IL-4 Antibody | 50 μL |
| RSP010-C04 | Streptavidin-HRP | 50 μL |
| RSP010-C05 | Washing Buffer (10×) | 50 mL |
| RSP010-C06 | Dilution Buffer (1×) | 50 mL |
| RSP010-C07 | AEC Dilution | 25 mL |
| RSP010-C08 | AEC Solution A (20×) | 0.8 mL |
| RSP010-C09 | AEC Solution B (20×) | 0.8 mL |
| RSP010-C10 | AEC Solution C (20×) | 0.8 mL |
This kit is used to detect mouse IL-4 at the cellular level.
It is for research use only.
2. Find the expiration date on the outside packaging and do not use reagents past their expiration date.
3. The opened kit should be stored per components table. The shelf life is 30 days from the date of opening.
Your experiment will include 6 simple steps:
a) A monoclonal antibody specific for mouse IL-4 has been pre-coated onto a polyvinylidene difluoride (PVDF)-backed microplate.
b) Appropriately stimulated cells are pipetted into the wells and the microplate is placed into a humidified 37 °C CO2 incubator for a specified period of time.During this incubation period, the immobilized antibody in the immediate vicinity of the secreting cells binds secreted IL-4.
c) After washing away any cells and unbound substances, a biotinylated antibody specific for mouse IL-4 is added to the wells. Following a wash to remove any unbound biotinylated antibody, HRP conjugated to streptavidin is added.
d) Wash the plate and add the Streptavidin-HRP diluted by Dilution Buffer to the plate.
e) Wash the plate and add AEC. A reddish-brown colored precipitate forms and appears as spots at the sites of cytokine localization, with each individual spot representing an individual IL-4 secreting cell.
f) The spots can be counted with an automated ELISpot reader system or manually using a stereomicroscope.
2.5×10⁵ cells/well Mouse Spleen Cells were stimulated with 18 μg/mL Positive Stimulus and cultured at 37℃ and 5%CO2 for 20h, 228 spots were produced. The following example data is for reference only.
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