Components
Pre-coated 96 well plate
Standard
Sample Diluent
Assay Buffer concentrate
Biotinylated Detection Antibody
SAV-HRP
Wash Buffer
Controls
Chromogen
Stop Solution
Adhesive Plate Covers
Material not included
Precision pipette, disposable 5-1000µL plastic tip, 5-15m plastic pipette
A two-liter glass or plastic container to prepare wash buffer
Automatic 96-well dish washer
1.5mL polypropylene or polyethylene tube prep standard
Disposable reagent reservoir
15mL plastic tube to prepare streptavidin-hrp solution
Standard ELISA reader for absorbance measurements at 450nm and 550nm.
Graphics paper or computerized curve fitting statistical software package
Assay Time
Less than 4.5 hours
Reagent Preparation
1. Wash Buffer
Wash buffers must be at room temperature before use in assays. If it becomes apparent, do not wash with water that has been contaminated during storage.
When using partial plates, store the reconstituted wash buffer at 2-8°C.
2. Standards
The lyophilized standard was reconstituted and used, per vial. Prepare standards before use and use within one hour of reconstitution. Do not store refactored criteria.
Assay Procedure
1. Add Incubation Buffer to all wells except the chromogen blank.
2. Add the standard, buffer solution, or cell culture medium sample to the appropriate wells. for
Serum and control samples, add standard dilution buffer, then add samples to
suitable well. Leave the chromogen blank holes blank.
3. Tap the side of the plate to mix. Cover the plate with a plate lid and incubate at room temperature for 2-3 hours.
4. Aspirate the solution thoroughly and wash the wells 4 times with wash buffer.
5. Add Biotin Conjugate solution to each well except the chromogen blank.
6. Cover the plate and incubate at room temperature for 1-2 hours.
7. Aspirate the solution thoroughly and wash the wells 4 times with wash buffer.
8. Add Streptavidin-HRP solution to each well except the chromogen blank.
9. Cover the plate with a plate cover and incubate at room temperature.
10. Aspirate the wells thoroughly and wash the wells 4-6 times with wash buffer.
11. Add stable chromogen to each well. The substrate solution begins to turn blue.
12. Incubate at room temperature for 30-45 minutes in the dark.
13. Add stop solution to each well. Tap the side of the plate to mix. The solution in the well changed from blue to yellow.
Calculation of Results
Calculate results using graph paper or curve-fitting statistical software. The human IL-6 amount in each sample is determined by interpolating from the absorbance value (Y axis) to human IL-6 concentration (X axis) using the standard curve.
Assay Precision
10.24-400 pg/mL
Format
Sandwich ELISA Kit
Precaution of Use
All products are supplied for research and laboratory use only.
Handling Advice
All blood components and biological materials should be treated with precautions as potentially hazardous. Strictly follow the management principles of the Centers for Disease Control and Prevention, Occupational Safety and Health Administration for handling and disposing of infectious agents.
Storage
The ELISA Kits are shipped at 2 to 8°C. Upon receipt, store the kits at 2 to 8°C in dark.
Expiry Date
Stability If properly stored, all components are stable for up to 12 months. For expiry dates for the entire kit, see the kit label. The expiration date of each ingredient is shown on the bottle label. The expiration date of a kit ingredient is guaranteed only if the ingredient is properly stored and, in the event of repeated use of an component, the reagent will not be contaminated by the first treatment.
Note
Each production lot of this ELISA kit is quality tested to meet criteria such as sensitivity, specificity, precision and lot-to-lot consistency. See the manual for more information on validation.
