The kinetics of most proteins involved in DNA damage sensing, signaling and repair following ionizing radiation
exposure cannot be quantified by current live cell fluorescence microscopy methods. This is because most of these
proteins, with only few notable exceptions, do not attach in large numbers at DNA damage sites to form easily detectable
foci in microscopy images. As a result a high fluorescence background from freely moving and immobile fluorescent
proteins in the nucleus masks the aggregation of proteins at sparse DNA damage sites. Currently, the kinetics of these
repair proteins are studied by laser-induced damage and Fluorescence Recovery After Photobleaching that rely on the
detectability of high fluorescence intensity spots of clustered DNA damage. We report on the use of Number and
Brightness (N&B) analysis methods as a means to monitor kinetics of DNA repair proteins during sparse DNA damage
created by γ-irradiation, which is more relevant to cancer treatment than laser-induced clustered damage. We use two
key double strand break repair proteins, namely Ku 70/80 and the DNA-dependent protein kinase catalytic subunit
(DNA-PK<sub>CS</sub>), as specific examples to showcase the feasibility of the proposed methods to quantify dose-dependent
kinetics for DNA repair proteins after exposure to γ-rays.
Several nonlinear effects (i.e., continuum generation, self-focusing, and material damage) were studied during femtosecond photodisruption. Numerical aperture dependence of white-light continuum generation and material damage were determined and a relation between the two effects was shown. We showed the possibility of reducing nonlinear side effects and at the same time ensuring precise cut by using lenses of a suitable numerical aperture for refractive surgery, cell surgery, and tissue dissection. Other side effects associated with optical breakdown in model substance were also discussed.