Components
Pre-coated 96 well plate
Standard
Sample Diluent
Assay Buffer concentrate
Biotinylated Detection Antibody
Amplification Diluent Concentrate
Amplification Reagent I
Amplification Reagent II
SAV-HRP
Wash Buffer
Chromogen
Stop Solution
Adhesive Plate Covers
Material not included
5 mL and 10 mL graduated pipettes
Adjustable single channel micropipette from 5 µL to 1000 µL with disposable tip
Adjustable multi-channel micropipette from 50 µL to 300 µL with disposable tip
Multichannel micropipette storage tank
Beakers, flasks and cylinders required for preparation of reagents
Washing liquid conveying device
Able to operate at 450 nm (620 nm as Optional reference wavelength)
Glass distilled water or deionized water
A statistical calculator with a linear regression program
Analysis.
Assay Time
Less than 3.5 hours
Reagent Preparation
1. The buffer concentrate should be brought to room temperature and diluted before starting the test procedure.
2. If crystals form in the buffer concentrate, heat gently until the crystals are completely dissolved. See the kit manual for the rest.
Assay Procedure
1. For the standard curve, add the standard to the corresponding hole. For sample, add diluted sample to the well.
2. Cover well and incubate at room temperature for 2-4 hours or gently shake at 4°C overnight.
3. Discard the solution and wash it 3-5 times with 1X wash buffer. Use a multi-channel pipette or automatic cleaner to fill each hole with a cleaning buffer. Complete liquid removal at each step is essential for good performance. After the last wash, remove any remaining wash buffer by suction or decanting. Turn the board upside down and drain with a clean paper towel.
4. Add the prepared biotin conjugates into each well.
5. Incubate gently at room temperature for 1-2 hour.
6. Discard the solution. Repeat the washing in Step 3.
7. Add the prepared Streptavidin-HRP solution to each well.
8. Incubate gently at room temperature, shaking for 30-50 minutes.
9. Discard the solution. Repeat the washing in Step 3.
10. Add TMB substrate to each well. The substrate will begin to turn blue.
11. Incubate at room temperature, dark, for 30-45 minutes, shaking gently.
12. Add termination solution to each well. Tap lightly on the sides of the boards to blend. The solution in the hole changes from blue to yellow.
Calculation of Results
1.Calculate the mean absorbance values for each set of replicate standards and samples.
2. Create a standard curve by plotting the mean absorbance of each standard concentration on the ordinate and the monkey IL-2 concentration on the abscissa.
3. To determine the circulating monkey IL-2 concentration for each sample, first find the mean absorbance value on the ordinate and extend a horizontal line onto the standard curve.
4. At the intersection, the vertical line is extended to the abscissa and the corresponding monkey IL-2 concentration is read.
5. The sample has been diluted, so the concentration read from the standard curve must be multiplied by the dilution factor.
Assay Precision
18.8-1,200 U/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.
