In Vitro Alpha Particle Irradiation: Methodological Determinants of Experimental Reliability

To develop and validate an easily implementable and low-cost in vitro alpha-particle irradiator that does not rely on specialized infrastructure.

We constructed an alpha particle irradiator composed of an ²⁴¹Am source and a specialized collimator. We also constructed a custom mylar-bottom dish ("alpha well") to allow alpha particle irradiation of cells cultured on the mylar surface. Monte Carlo simulations, gafchromic films and nuclear track detectors were used to characterize the dose rate and linear energy transfer (LET) spectrum at the surface of the alpha well where cells reside. Clonogenic cell survival and DNA damage across several cancer cell lines were used to optimize and validate the irradiation workflow.

Gafchromic film measurements showed a homogenous irradiation field across the surface of the alpha well. Monte Carlo simulations of the detailed setup of the irradiator determined a dose rate of 3.4 mGy/s and a dose-averaged LET in water of 111 keV/μm at the surface of the alpha well. Within their experimental limitations, the nuclear track detectors provided an additional confirmation of the Monte Carlo simulations. We showed that cell clustering and unattached cells can impair homogeneous traversal of alpha particles through the irradiated cell population, creating a mixed population of irradiated and non-irradiated cells. This limitation was minimized by using fibronectin coating to promote monolayer attachment and optimized washing steps to remove unattached cells. Alpha particles induced greater cell killing than x-rays, with relative biological effectiveness values generally above 2, and increased persistent γH2AX foci frequency and larger foci volumes at 48 h after irradiation, consistent with more clustered DNA damage distributed along particle tracks relative to x-rays.

This work provides a reproducible methodologic framework for in vitro alpha particle studies and establishes a practical platform suitable for investigating the biological mechanisms of alpha particles.