Human hypoglycosylated IgA1 (UGIGA1) ELISA kit experimental principle - Database & Sql Blog Articles

Human hypoglycosylated IgA1

(

UGIGA1

)

ELISA

Kit

Experimental Principle

This kit is designed for research purposes only.

Experimental Principle

The ELISA Kit uses a double-antibody sandwich assay to detect human hypoglycosylated IgA1 in biological samples. A purified monoclonal antibody specific to hypoglycosylated IgA1 (UGIGA1) is coated onto a microplate, and the sample is added to the wells. The target antigen binds to the immobilized antibody. An HRP-labeled secondary antibody then recognizes the bound antigen, forming an antibody-antigen-enzyme complex. After washing, TMB substrate is added, and the reaction is stopped with a stop solution. The intensity of the color produced correlates with the concentration of UGIGA1 in the sample. The absorbance is measured at 450 nm using a microplate reader, and the concentration is determined by comparing it to a standard curve.

Human hypoglycosylated IgA1

(

UGIGA1

)

ELISA

Kit

Composition

1. 130× Washing Solution: 20ml × 1 bottle

2. Stop Solution: 6ml × 1 bottle

3. Enzyme Standard Reagent: 6ml × 1 bottle

4. Standard (16μg/ml): 0.5ml × 1 bottle

5. Enzyme-Labeled Coating Plate: 12×8

6. Sample Diluent: 6ml × 1 bottle

7. TMB Substrate A: 6ml × 1 bottle

8. TMB Substrate B: 6ml × 1 bottle

9. Standard Dilutions: 1.5ml × 1 bottle

10. Instructions: 1 copy

11. Sealing Film: 2 sheets

12. Seal Bag: 1 piece

Sample Requirements

1. Samples should be processed as soon as possible after collection. If not tested immediately, store at -20°C, avoiding repeated freeze-thaw cycles.

2. Avoid samples containing NaN3, as it may inhibit HRP activity.

Human hypoglycosylated IgA1

(

UGIGA1

)

ELISA

Kit

Procedure

1. Standard Dilution: Prepare a series of standards from the original stock.

2. Sample Loading: Load 50μl of standard and 50μl of sample diluent plus 10μl of sample (final 5× dilution).

3. Incubation: Seal and incubate at 37°C for 30 minutes.

4. Washing: Wash 5 times with diluted washing buffer.

5. Add Enzyme: Add 50μl of enzyme-labeled reagent to each well except blank.

6. Incubation: Repeat incubation at 37°C for 30 minutes.

7. Washing: Repeat washing steps.

8. Color Development: Add 50μl of TMB A and B, incubate for 10 minutes at 37°C.

9. Stop Reaction: Add 50μl of stop solution to each well.

10. Read OD: Measure absorbance at 450 nm within 15 minutes.

Calculation

Create a standard curve using known concentrations and OD values. Calculate the unknown sample concentration based on the standard curve or linear regression equation. Multiply by the dilution factor to obtain the actual concentration.

Human hypoglycosylated IgA1

(

UGIGA1

)

ELISA

Kit

Precautions

1. Allow the kit to reach room temperature before use. Store unused enzyme-labeled components in a sealed bag.

2. If washing solution crystallizes, warm it gently before use.

3. Use accurate pipetting and avoid cross-contamination. For large numbers of samples, use a multichannel pipette.

4. Always run a standard curve, preferably in duplicate. If sample OD exceeds the highest standard, dilute the sample and adjust the calculation accordingly.

5. Use one sealing film per plate to prevent contamination.

6. Keep substrates away from light.

7. Follow the manual strictly and verify results with a microplate reader.

8. Treat all waste materials as biohazardous.

9. Do not mix reagents from different batches.

Storage Conditions and Expiration

1. Store at 2–8°C.

2. Shelf life: 6 months from date of manufacture.