Assay Type
Solid Phase Sandwich ELISA
Format
96-well strip plate
Assay Length
4.5 hours
Sample Type & Volume Required Per Well
Cell Culture Supernates (100 uL), Serum (100 uL), EDTA Plasma (100 uL), Heparin Plasma (100 uL), Citrate Plasma (100 uL)
Sensitivity
0.7 pg/mL
Assay Range
3.1 - 300 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma, Citrate Plasma)
Specificity
Natural and recombinant human IL-6
Cross-reactivity
< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
Interference
No significant interference observed with available related molecules.
Control Available
QC01-1, Quantikine Immunoassay Control Group 1 - Please InquireProduct Summary
The Quantikine Human IL-6 Immunoassay is a 4.5 hour solid phase ELISA designed to measure IL-6 in cell culture supernates, serum, and plasma. It contains recombinant human IL-6 and antibodies raised against recombinant human IL-6 and has been shown to accurately quantitate the recombinant factor. Results obtained using natural IL-6 showed linear curves that were parallel to the standard curves obtained using the E. coli-expressed Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for natural human IL-6.
Precision
Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision.
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in separate assays to assess inter-assay precision.
Cell Culture Supernates
| Intra-Assay Precision | Inter-Assay Precision |
|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean | 15.8 | 95.6 | 179 | 16.4 | 98.8 | 188 |
| Standard Deviation | 0.7 | 3 | 3.1 | 0.6 | 2.5 | 3.7 |
| CV% | 4.4 | 3.1 | 1.7 | 3.7 | 2.5 | 2 |
Serum, EDTA Plasma, Heparin Plasma, Citrate Plasma
| Intra-Assay Precision | Inter-Assay Precision |
|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean | 16.8 | 97.7 | 186 | 17.2 | 101 | 191 |
| Standard Deviation | 0.7 | 1.6 | 3.8 | 1.1 | 3.3 | 7.2 |
| CV% | 4.2 | 1.6 | 2 | 6.4 | 3.3 | 3.8 |
Recovery
The recovery of human IL-6 spiked to three different levels throughout the range of the assay in various matrices was evaluated.
| Sample Type | Average % Recovery | Range % |
|---|
| Cell Culture Media (n=5) | 98 | 94-103 |
| Citrate Plasma (n=5) | 91 | 82-95 |
| EDTA Plasma (n=5) | 95 | 84-101 |
| Heparin Plasma (n=5) | 90 | 88-98 |
| Serum (n=5) | 93 | 86-99 |
Linearity
To assess the linearity of the assay, four samples were spiked with high concentrations of IL-6 in various matrices and diluted with the appropriate Calibrator Diluent to produce samples with values within the dynamic range of the assay.
Preparation and Storage
Refer to the product for complete assay procedure.
- Prepare all reagents, standard dilutions, and samples as directed in the product insert.
- Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.
100 μL Assay Diluent
- Add 100 μL of Assay Diluent to each well.
100 μL Standard, Control, or Sample - Add 100 μL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.
- Aspirate each well and wash, repeating the process 3 times for a total of 4 washes.
200 μL Conjugate - Add 200 μL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.
- Aspirate and wash 4 times.
200 μL Substrate Solution
- Add 200 μL Substrate Solution to each well. Incubate at room temperature for 20 minutes. PROTECT FROM LIGHT.
50 μL Stop Solution
- Add 50 μL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.
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