TECHNOLOGY

Q1: What is the role of plate washing?​​

To remove unbound antigens, antibodies, and other impurities, reduce non-specific reactions, and improve detection sensitivity, specificity, and accuracy.

 

Q2: How to prepare the wash buffer?​​

The kit-supplied wash buffer is a 30 mL, 25X concentrated solution, sufficient for up to 96-well plates. For example, if running 48 wells (standard + samples):

•​Concentrated buffer needed: 30mL×48/96=15mL

•​Dilution: Mix 15 mL of 25X concentrate with 15×24=360mL sterile distilled water.

 

Q3: What are common plate washing methods?​​

Manual plate washing and automated plate washer.

 

Q4: How much wash buffer per well?​​

The working volume of a general ELISA experiment does not exceed 200μL, while the single-well volume of most microplate readers is around 350μ L. To ensure the washing effect, the washing solution needs to exceed the working volume. The washing solution for ELISA experiments is generally 300μL/well.

 

Q5: What happens if washing is insufficient?​​

​Symptoms: High background signal (false positives), non-specific binding residue ("all-blue plate").

​Cause: Unbound antibodies or antigens have not been fully removed.

​Solution: Follow the recommended wash times according to the datasheet.

 

Q6: What are the risks of over-washing?​​

Excessive washing of the plate can wash away the already bound antigen-antibody complexes, leading to a weakened overall color development and even a "white board". Therefore, the plates should be washed strictly in accordance with the number of times specified in the datasheet.

 

Q7: Which operations cause cross-contamination between wells?​​

1. Inserting pipette tips too deep into the well or touching the bottom.
2. Overflowing wash buffer onto adjacent wells. High-precision pipettes and pipette tips should be selected. Before use, it is also necessary to check to ensure that the pipette tips are installed tightly.
3. Incorrect plate slapping technique. The correct way is to apply force with the wrist and quickly flip and spin dry.
4. Repeatedly tapping the same spot on absorbent paper. The correct approach should be that after taking a pat, if there are visible water stains on the absorbent paper, a new absorbent paper should be replaced immediately.

Q8: Prepare next-step reagents before or after plate washing?​​

Prepare reagents before washing to avoid drying and loss of reagent activity.

 

Q9: What are the requirements for absorbent paper?​​

Use lint-free, dust-free paper with good absorbency. Avoid repetitive tapping in the same spot.

 

Q10: How many taps are needed to dry the plate?​​

3–5 taps until no visible moisture remains on absorbent paper.

 

Q11: What is the components of wash buffer?​​

Typically PBS (pH 7.4) or TBS with 0.05%–0.5% Tween-20/Triton X-100. Some formulations include BSA/NaN₃ or NaCl for ionic strength. Note: Our wash buffer does not contain BSA/NaN₃.

 

Q12: Can different manufacturers’ wash buffers be mixed or can the same manufacturers’ different wash buffers be mixed?​​

No. Incompatible experiment system and formulations may disrupt assay performance.

 

Q13: Are precipitates in concentrated wash buffer normal?​​

As is a 25X concentrated washing solution with a relatively high concentration, due to long-term storage or temperature changes, there will be a certain degree of crystallization and precipitation, which is a normal phenomenon, gently shaking or in a 37℃ water bath to dissolve before use.