Why Doesn't Two-Photon Microscopy Damage Samples Everywhere?

[Ben Hom Chen]

I know two-photon microscopy use the longer wavelength laser,but with more photon density.To my knowledge,the laser with more photon density means it is very strong(High Power?).But articles always say that two-photon microscopy only damage on focus point and is suitable to investigate biologic sample,while one-photon confocal microscopy will damage the sample everywhere that light can reach.

But I really wonder,if two-photon microscopy have a stronger laser(more photon number in short time),why wouldn't it damage the sample everywhere?

Is the laser harmful only after the sample is excited?

Thx>.<

 

[Valerie Park]

Yes, the laser used in two-photon microscopy is generally more powerful than the laser used in one-photon confocal microscopy. However, only the area of the sample that is in focus will be excited and damaged by the laser. This is because two-photon microscopy uses a longer wavelength laser which requires two photons to be absorbed at the same time by the sample in order to be excited. This means that the laser needs to be highly focused on the specific area of the sample that needs to be examined, otherwise the laser will not be able to excite the sample. In contrast, one-photon microscopy uses a shorter wavelength laser which only requires one photon to be absorbed in order to be excited, so it is not as focused as two-photon microscopy and can therefore damage the sample even outside of the intended area of focus.

 

[astrokreng]

Two-photon microscopy is a powerful imaging technique that utilizes a longer wavelength laser with a higher photon density compared to one-photon confocal microscopy. This allows for better penetration and less scattering in biological samples, making it a valuable tool for investigating biological structures and processes.

It is true that the higher photon density of two-photon microscopy means that the laser is stronger, but this does not necessarily mean it is harmful to the sample. The key difference between two-photon and one-photon microscopy is the way in which the laser interacts with the sample.

In one-photon microscopy, the laser directly excites the sample at the focus point, causing potential damage to the surrounding areas as the laser moves across the sample. However, in two-photon microscopy, the photons are absorbed in pairs, only at the focal point, resulting in a more precise and localized excitation. This means that the laser is only harmful to the sample at the specific focus point and not throughout the entire sample.

Additionally, the laser is not harmful until it reaches the sample and is absorbed by the molecules. The intensity of the laser is carefully controlled to ensure that it does not cause any damage to the sample until it reaches the focus point. This is why two-photon microscopy is considered to be a safer and gentler imaging technique for biological samples.

In summary, the higher photon density of two-photon microscopy does not necessarily mean it is more harmful to the sample. The unique way in which the laser interacts with the sample and the careful control of laser intensity ensure that it is only harmful at the focus point, making it a valuable tool for studying biological samples.