Two-photon microscope captures plant cells
Leiden physicists are helping Wageningen plant researchers to study unpredictable plant embryos. For this, they are using a novel two-photon fluorescence microscope, aided by a 30 thousand euro ZonMW grant.
Any scientific instrument, however brilliant, can be improved. Take, for example, the fluorescence microscope, which is very useful when one wants to track a biochemical process up close.
The proteins within the cell that have your interest, can be coupled to a special fluorescent protein from luminescent jellyfish: GFP. When green light hits this protein, it will fluoresce. Electrons within the GFP protein jump one energy level higher, and fall back immediately. In doing so, they emit green light with a slightly higher wavelength than the original incoming light. By imaging this returning light, the GFP within the cell is accurately visible, and with that, the interesting molecule which it's coupled to is visible as well.
Improvements are possible
Of course, this technique can be improved upon, explains physicist John van Noort. 'We use infrared light instead of green light'. The energy carried by one green photon needed to bump up the electron can also be delivered by two infrared photons absorbed at the same time.
This two-photon technique has a number of advantages, Van Noort explains. Infrared light penetrates deeper into tissues, so the view is deeper. The image resolution is higher: within the focus of the infrared laser beam, only a small volume has enough photons for the two-photon excitation.
In general, this yields images that differ greatly from the ordinary fluorescence microscopy images, the researchers noticed when studying plant cells. Only the two-photon images show how the protein LEC-is concentrated within small vesicles around the plant cell nucleus of Brassica napus (rapeseed).
This is interesting in connection with the development of plant embryos from adult planst. 'Specific chemical compounds can turn adult plant cells back into plant embryos', says Van Noort. This process is a very common multiplication method for plant breeders.
The LEC-1 protein is one of the first markers showing that this change has taken place, but a full understanding of this process is still missing, says Van Noort. 'It works for one cell, but for another cell, it inexplicably doesn't.'
Together with researcher Kim Boutilier at Wageningen University & Research and plant breeding company KWS Saat the Leiden physicists will investigate this process, thanks to a 30 thousand euro research grant from ZonMW.
This way, we make available the microscopy infrastructure in the Leiden Cell Observatory for researchers in the entire country, Van Noort explains. 'We have the microscope, they have the research questions.'