BETAview: a digital β-imaging system for dynamic studies of biological phenomena

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Abstract

We present a digital autoradiography (DAR) system, named BETAview, based on semiconductor pixel detectors and a single particle counting chip, for quantitative analysis of β-emitting radioactive tracers in biological samples. The system is able to perform a real time monitoring of time-dependent biological phenomena. BETAview could be equipped either with GaAs or with Si semiconductor pixellated detectors. In this paper, we describe the results obtained with an assembly based on a Si detector, 300 μm thick, segmented into 64×64 170 μm size square pixels. The detector is bump-bonded to the low threshold, single particle counting chip named Medipix1, developed by a CERN-based European collaboration. The sensitive area is about 1 cm2. Studies of background noise and detection efficiency have been performed. Moreover, time-resolved cellular uptake studies with radiolabelled molecules have been monitored. Specifically, we have followed in vivo and in real time, the [14C]l-leucine amino acid uptake by eggs of Octopus vulgaris confirming the preliminary results of a previous paper. This opens the field of biomolecular kynetic studies with this new class of semiconductor DAR systems, whose evolution (using the Medipix2 chip, 256×256 pixels, 55 μm pixel size) is soon to come.

Introduction

Autoradiography (AR) is a widespread and powerful analysis tool in the medical and biological fields for applications ranging from tissue imaging and DNA sequencing to molecular metabolic path reconstruction.

Traditional AR is still the commonly used technique and is based on photographic film, which features excellent spatial resolution over large area. However, it is affected by non-linear response at low activity, low detection efficiency and high level of noise. Digital Autoradiography (DAR), based on detection systems able to convert the ionization radiation into electrical signals, can be thought of as the natural evolution of conventional AR: the data acquisition, the storage and the display of the detected radiation is performed by digital electronics. Examples of DAR systems are those based on storage phosphor plates [1] or on MWPC [2], [3], or on silicon microstrips detector [4], [5].

We have developed a new DAR system (BETAview), based on a single particle counting solid-state detector, which offers several advantages such as: “unlimited” (in principle) linear dynamic range, linearity, good sensitivity at low activity, no developing time and real time imaging, capability of following the dynamic evolution of a biological process, possibility of using multiple markers and of acquiring differential or integrated images. In previous papers [6], [7], we presented some preliminary experiments with GaAs detectors, in order to show its potential in different biological applications not requiring large sensitive areas, due to the limited dimension of the present sensor (about 1 cm2).

In particular, in order to test the capability of the device to follow a biological process where low-activity (⪡1 cps/cm2) and low-energy β(<150 keV) markers are used, we performed an experiment on Octopus vulgaris eggs. Octopus vulgaris eggs consist of an oocyte surrounded by a monolayer of follicle cells. Their planar dimensions are about 0.5×1 mm2, thus being well suited to be imaged with our system which has a matrix array of 64×64 pixels, each of size 0.17×0.17 mm2.

In the experiment, reported here, we monitor, on-line, a 14C-labelled amino acid uptake by the Octopus eggs living in the culture medium. Specifically, we want to observe the accumulation of radioactivity into single-egg cells, after putting the marked amino acid in the surrounding medium. This would enable to study, in vivo, the kinetics of biomolecules in a biological system.

Section snippets

System description

BETAview is based on a solid-state pixel array detector, bump-bonded pixel to pixel to a low-threshold, single-particle counting electronics, the photon counting chip (PCC) developed by the Medipix1 Collaboration for medical applications [8]. Each cell (corresponding to a detector pixel) of the front-end chip has a maximum count rate of 2 MHz, a minimum nominal threshold of 1400 e (corresponding to 5.1 keV in Si) and contains a 15-bit counter. Detectors used are 300 μm thick Si 64×64 pixel

Biological sample preparation

The Octopus vulgaris eggs were removed from ovary and incubated in a solution of seawater and a nutritive medium. Each egg at this stage consists of an oocyte surrounded by an epithelium of several hundred follicle cells, which are the cells internal to the egg where protein synthesis occurs. Eggs were 1–2 mm long and ≈0.5 mm wide. An environment rich in [14C]l-leucine, the most representative amino acid in proteins, was prepared for the eggs: 200 μl nutritive medium, 2 μl [14C]l-leucine (1.85×106

Conclusion

This experiment shows the realistic capability of the BETAview system for dynamic autoradiographic studies. For this purpose, we have followed with success, in real time, the kinetics of the uptake of an amino acid in a biological system. The high sensitivity of the system was also demonstrated, since the estimated activity in each egg was less than 1 Bq. With the 300 μm silicon detector, the noise level of the system is 3.5×10−2 cps/cm2 and the minimum detectable activity is 0.3 Bq/cm2 (15 keV

Acknowledgements

The authors thank Drs. A. Di Cosmo and C. Di Cristo (Department Of Zoology, University of Napoli Federico II) for providing the Octopus Vulgaris eggs and their culture medium.

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