Plant folic acid (FA) ELISA kit operating instructions - Database & Sql Blog Articles

Plant Folic Acid (FA) ELISA Kit

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

Experimental Principle:

The Plant Folic Acid (FA) ELISA Kit utilizes a double-antibody sandwich assay to determine the concentration of folic acid in plant samples. The microplate is pre-coated with purified anti-FA antibodies, which bind to FA present in the sample. After incubation, HRP-labeled anti-FA antibodies are added, forming an antibody-antigen-enzyme-labeled antibody complex. Following washing steps, TMB substrate is introduced, which turns blue under the action of HRP and then changes to yellow when an acidic stop solution is added. The intensity of the color is directly proportional to the FA concentration in the sample. Absorbance at 450 nm is measured using a microplate reader, and the FA concentration is calculated from a standard curve.

Kit Components:

1. 30× Washing Solution: 20ml × 1 bottle

2. Enzyme Standard Reagent: 6ml × 1 bottle

3. Enzyme-Labeled Coating Plate: 12 wells × 8 strips

4. Sample Diluent: 6ml × 1 bottle

5. Reagent A: 6ml × 1 bottle

6. Color Developer B: 6ml × 1 bottle

7. Stop Solution: 6ml × 1 bottle

8. Standard (24μg/L): 0.5ml × 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 extracted as soon as possible after collection and tested promptly. If testing is delayed, store samples at -20°C, avoiding repeated freeze-thaw cycles.

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

Procedure:

1. Standard Dilution: Prepare a serial dilution of the original standard according to the provided chart.

2. Loading: Add 50 μl of standard, 40 μl of sample diluent, and 10 μl of sample into each well. Mix gently.

3. Incubate at 37°C for 30 minutes.

4. Wash 5 times with 30× diluted washing solution.

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

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

7. Wash again 5 times.

8. Add 50 μl of TMB developer to each well and incubate at 37°C for 15 minutes.

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

10. Measure OD values at 450 nm within 15 minutes of adding the stop solution.

Calculation:

Plot a standard curve using standard concentrations and corresponding OD values. Determine the sample concentration by interpolation or linear regression. Multiply the result by the dilution factor to obtain the actual sample concentration.

Precautions:

1. Allow the kit to reach room temperature before use. Store unopened enzyme reagents in a sealed bag.

2. If the washing solution crystallizes, warm it in a water bath before use.

3. Use accurate pipettes and ensure consistent timing during loading.

4. Always prepare a standard curve and run duplicate wells. If sample OD exceeds that of the first standard, dilute the sample before testing.

5. Use a new sealing film for each experiment to prevent contamination.

6. Keep the substrate away from light.

7. Follow the manual strictly and rely on microplate reader results.

8. Treat all waste materials as biohazardous.

9. Do not mix components from different batches.

10. In case of discrepancies, the English version of the manual takes precedence.

Storage Conditions and Expiration:

1. Store the kit at 2–8°C.

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