Direct observation of unstained wet biological samples by scanning-electron generation X-ray microscopy

Analytical tools of nanometre-scale resolution are indispensable in the fields of biology physics and, chemistry One suitable tool, the soft X-ray microscope, provides high spatial resolution of visible light for wet specimens For biological specimens, X-rays of water-window wavelength between carbon (284 eV; 4.3 nm) and oxygen (540 eV, 2.3 nm) absorption edges provide high-contrast imaging of biological samples in water. Among types of X-ray microscope, the transmission X-ray microscope using a synchrotron radiation Source with diffractive zone plates offers the highest spatial resolution. approaching 15-10 nm. However even higher resolution is required to measure proteins and protein complexes in biological specimens: therefore, a new type of X-ray microscope with higher resolution that uses a simple light source is desirable. Here we report a novel scanning-electron generation X-ray microscope (SGXM) that demonstrates direct imaging of unstained wet biological specimens. We deposited wet yeasts in the spice between two silicon nitride (Si3N4) films. A scanning electron beam of accelerating voltage 5 keV and current 1.6 nA irradiates the titanium (Ti)-coated Si3N4 film, and the soft X-ray signal from it is detected by an X-ray photodiode (PD) placed below the sample The SGXM can theoretically achieve better than 5 nm resolution Our method call be utilized easily for various wet biological samples of bacteria. viruses, and protein complexes. (c) 2009 Elsevier Inc. All rights reserved