Human hypoglycosylated IgA1 (UGIGA1) elisa detection kit instructions - Database & Sql Blog Articles

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Kaixin micro test

Test probe P100-M3

Brand AVX TPSE226M035R0125 Low impedance tantalum capacitor AVX 22

Human hypoglycosylated IgA1 (UGIGA1)

ELISA Test

Kit Instructions

This kit is for research use only and not intended for diagnostic or therapeutic purposes.

Experimental Principle

The ELISA kit uses a double-antibody sandwich assay to detect the concentration of human hypoglycosylated IgA1 (UGIGA1) in the sample. A microplate is pre-coated with a purified anti-UGIGA1 antibody, forming a solid-phase antibody. The sample containing low glycosylated IgA1 is added to the coated wells, followed by incubation with HRP-labeled anti-UGIGA1 antibody. This forms an antibody-antigen-enzyme-labeled antibody complex. After thorough washing, the substrate TMB is added, which turns blue under the catalysis of HRP and then changes to yellow upon acid addition. The intensity of the color is directly proportional to the amount of UGIGA1 in the sample. The optical density (OD) is measured at 450 nm using a microplate reader, and the concentration of UGIGA1 is calculated from a standard curve.

Human Hypoglycosylated IgA1 (UGIGA1)

ELISA Test

Kit Composition

1. 130x 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 wells × 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. Instruction Manual – 1 copy

11. Sealing Film – 2 sheets

12. Sealed Bag – 1

Sample Requirements

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

2. Samples containing NaN3 cannot be used, as it may inhibit HRP activity.

Human Hypoglycosylated IgA1 (UGIGA1)

ELISA Test

Kit Steps

1. Standard Dilution: The original standard can be diluted according to the provided dilution chart. For example:

• Add 150 μl of original standard to 150 μl of standard diluent to get 8 μg/ml (No. 5)

• Add 150 μl of No. 5 to 150 μl of diluent to get 4 μg/ml (No. 4)

• Continue this process to create standards down to 0.5 μg/ml (No. 1).

2. Loading: Set up blank, standard, and sample wells. Load 50 μl of standard and 40 μl of sample diluent into each well, then add 10 μl of sample. Mix gently.

3. Incubate at 37°C for 30 minutes.

4. Wash the plate 5 times using 30x diluted washing solution (diluted with distilled water).

5. Add 50 μl of enzyme reagent to all wells except the blank.

6. Incubate again at 37°C for 30 minutes.

7. Wash the plate again as before.

8. Add 50 μl of TMB A and 50 μl of TMB B to each well. Incubate at 37°C for 10 minutes.

9. Add 50 μl of stop solution to each well to terminate the reaction.

10. Measure OD at 450 nm within 15 minutes after stopping the reaction.

Calculation

Plot the standard curve using standard concentrations vs. OD values. Determine the sample concentration based on the OD value from the curve, then multiply by the dilution factor. Alternatively, use linear regression to calculate the sample concentration from the equation derived from the standard curve.

Human Hypoglycosylated IgA1 (UGIGA1)

ELISA Test

Kit Precautions

1. Allow the kit to reach room temperature for 15–30 minutes before use. Store unopened enzyme reagents in a sealed bag.

2. If the washing solution crystallizes, heat it in a water bath before use. It will not affect results.

3. Use a pipette for accuracy. Ensure proper calibration and keep loading time under 5 minutes. For large batches, consider using an automated dispenser.

4. Always prepare a standard curve and run duplicates. If the sample OD is higher than the highest standard, dilute the sample before testing and adjust the calculation accordingly.

5. Use a new sealing film for each experiment to avoid cross-contamination.

6. Keep the substrate away from light during storage and use.

7. Strictly follow the protocol to ensure accurate results.

8. All samples, washes, and waste should be treated as biohazardous materials.

9. Do not mix components from different batches.

Storage Conditions and Expiration Date

1. Store the kit at 2–8°C.

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