If anyone can suggest for about APEX2-Gold methods ?

12 January 2023 1 3K Report

Recently, I established a mutant strain of Drosophila with an APEX2 tag in its genome. Using this strain, I have successfully performed normal immunoelectron microscopy. However, it is difficult to detect the weak signal (it can be observed by confocal microscopy). Therefore, I would like to try the APEX2-Gold method. what points should I pay attention to when performing the APEX2-Gold method on animals? Also, can I keep the enhancement solution in stock? I would appreciate any tips anyone can give me.

https://elifesciences.org/articles/64630

Mohammad Alzeyadi

Hello,

When performing the APEX2-Gold method on animals, there are a few things to consider:

Make sure that your sample is properly fixed and dehydrated before staining. This will ensure that the APEX2 antibody can bind to the APEX2 tag in your mutant strain.

Use a high-resolution electron microscope, such as a transmission electron microscope (TEM), to visualize the gold particles. This will allow you to see the fine details of the APEX2-tagged proteins in your sample.

Use a high-quality APEX2 antibody and gold conjugate. This will ensure that the signal is strong and specific to the APEX2 tag in your mutant strain.

The enhancement solution is usually not stable for long periods of time, it is best to prepare fresh for each experiment.

Optimize the conditions for the staining by testing different concentrations of the reagents and incubation times.

Overall, the APEX2-Gold method is a powerful tool for detecting and analyzing APEX2-tagged proteins in electron microscopy. With proper technique and high-quality reagents, you should be able to detect a strong signal in your samples.

Regards/

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