What does it mean that GFP has poor resistance to photobleaching

It is easy to be bleached by photo.

Green fluorescent protein, or GFP for short, was first discovered in 1962 by Shimomura et al. in a jellyfish with the scientific name Aequorea Victoria. The proteins produced by their genes emit green fluorescence when excited by light in the blue wavelength range. This luminescence process also requires the help of the cold light protein AEQUORIN, and this cold light protein can interact with calcium ions (Ca+2).

GFP fluorescence is extremely stable, and the anti-photobleaching ability of GFP is stronger than that of fluorescein under excitation light, especially at 450 490nm blue light wavelength. GFP needs to be fluorescent in the oxidized state, and a strong reducing agent can make GFP into a non-fluorescent form, but once reexposed to air or oxygen, GFP fluoresces.

GFP fluorescence is extremely stable, and under excitation light irradiation, GFP has stronger resistance to photobleaching than fluorescein, especially at 450 490nm blue wavelength.

GFP needs to fluoresce in an oxidized state, and a strong reducing agent converts GFP to a non-fluorescent form, but once reexposed to air or oxygen, GFP fluorescence is restored immediately. Some weak reducing agents do not affect GFP fluorescence. Moderate oxidants also have little effect on GFP fluorescence, such as the fixation of biological materials, the dehydrating agent glutaric acid or formaldehyde.

The fluorescence sensitivity of GFP-fusion proteins is much higher than that of fluorescein-labeled fluorescent antibodies, and the resistance to photobleaching is stronger, so it is more suitable for quantitative determination and analysis. However, because GFP is not an enzyme, the fluorescence signal is not enzymatically amplified, so GFP may be less sensitive than some enzyme reporter proteins.

Since GFP fluorescence is an autonomous function of biological cells, the production of fluorescence does not require any exogenous reaction substrate, so GFP, as a widely used living reporter protein, has an effect that is incomparable to any other enzyme reporter protein.

The plasmid can be copied and transfected to allow it to be expressed intracellularly, and it can fluoresce green under the excitation light of a mercury lamp. It is generally used as a reference in experiments. Here's a simple example:

The construction of interfering cell lines is that the expression of green fluorescent protein can be seen to interfere with the process of line construction. In addition, through immunological techniques, the protein expression is displayed, which is generally semi-quantitative.