**Detection Range:** 48T200 ng/L – 5000 ng/L
**Purpose:** This kit is designed to quantitatively determine the concentration of Interleukin-23 (IL-23) in human serum, plasma, and other related biological samples.
**Experimental Principle:**
The IL-23 ELISA Kit utilizes a double-antibody sandwich immunoassay method. A microplate is pre-coated with a specific monoclonal antibody against IL-23. After adding the sample, IL-23 binds to the immobilized antibody on the plate. An HRP-conjugated secondary antibody is then added, forming a complex of antibody-antigen-enzyme-labeled antibody. Following thorough washing, TMB substrate is introduced, producing a blue color that turns yellow in the presence of an acidic stop solution. The intensity of the color is directly proportional to the IL-23 concentration in the sample. The optical density (OD) at 450 nm is measured using a microplate reader, and the IL-23 concentration is determined by comparing the results to a standard curve.
**Kit Components:**
1. 20× Washing Solution – 20ml × 1 bottle
2. Stop Solution – 3ml × 1 bottle
3. Enzyme Standard Reagent – 3ml × 1 bottle
4. Sample Diluent – 3ml × 1 bottle
5. Color Reagent A – 3ml × 1 bottle
6. Color Reagent B – 3ml × 1 bottle
7. Enzyme-Labeled Coating Plate – 12 wells × 4
8. Standard Product (10000 ng/L) – 0.5ml × 1 bottle
9. Standard Dilutions – 1.5ml × 1 bottle
10. Instructions – 1 part
11. Sealing Film – 2 sheets
12. Seal Bag – 1 piece
**Sample Requirements:**
Samples should be processed as soon as possible after collection. Follow recommended protocols for extraction. If testing cannot be performed immediately, store samples at -20°C, avoiding repeated freeze-thaw cycles. Samples containing NaN3 are not suitable for testing, as it may inhibit HRP activity.
**Procedure:**
1. **Standard Dilution:** Prepare a series of standards from the original stock (10000 ng/L) by diluting according to the provided chart.
2. **Sample Addition:** Set up blank, standard, and sample wells. Add 50 µL of standard or 40 µL of sample diluent plus 10 µL of sample (final 5-fold dilution). Avoid touching the well walls during addition.
3. **Incubation:** Seal the plate and incubate at 37°C for 30 minutes.
4. **Washing:** Use 20× diluted washing solution. Wash 5 times, ensuring complete removal of unbound reagents.
5. **Enzyme Addition:** Add 50 µL of enzyme-labeled reagent to each well except the blank.
6. **Second Incubation:** Incubate again at 37°C for 30 minutes.
7. **Color Development:** Add 50 µL of TMB A and 50 µL of TMB B, mix gently, and incubate at 37°C for 15 minutes in the dark.
8. **Stop Reaction:** Add 50 µL of stop solution to each well to halt the reaction.
9. **Measurement:** Read OD values at 450 nm within 15 minutes of stopping the reaction.
This kit provides a reliable and accurate method for IL-23 detection, ideal for research and clinical applications. Always follow the instructions carefully to ensure consistent and reproducible results.
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