Direct digital imaging of radio‐labeled two‐dimensional gel beta emissions using micro‐channel plate image enhancement

Abstract

Linus ~a u~i n gInstitute of Science and gel beta emissions using micro-channel plate image enhancement* Medicine, Palo Alto, CAThe use of autoradiography to record the distribution of radio-labels such as tritium or 35S in two-dimensional (2-D) gels has well known deficiencies: the film requires long exposures, has a nonlinear response curve, and has a restricted dynamic range. Autofluorography can improve upon each of these problems but only at the cost of resolution and quantitative accuracy. This paper demonstrates that a micro-channel plate electron multiplier and a resistive anode position detector can detect and localize individual beta emissions from 35S or tritium with at least 200 pm resolution. The locations of these beta emissions can be collected, converted to a 2-D image, and displayed on a graphics terminal or integrated over spot areas for quantitative estimates of spot intensity. The collection process is rapid, linear, and has a dynamic range far greater than that of film. With the 35S-test gel (which required 40 h for an autoradiogram), strong spots can be integrated with better than ten percent accuracy with less than five minutes of data collection. Weak spots (those with less than 4 cpm) may require 80 min or more. Tritium gels require collection times an order of magnitude longer for equivalent quantitative accuracy.

Rapid, high-sensitivity imaging of radiolabeled gels with microchannel plate detectors

“…A review of the electrophoresis literature shows that our imaging modality is similar to earlier work published in two papers in the mid-nineteen eighties [10,11]. Burbeck et al [10] reported an MCP-based detector of 40 mm diameter having a spatial resolution (200 mm) much poorer than either of our current detectors [7].…”

“…Burbeck et al [10] reported an MCP-based detector of 40 mm diameter having a spatial resolution (200 mm) much poorer than either of our current detectors [7]. While their system was capable of imaging tritiated labeled gels, neither of their papers gave measurements of the efficiency or linearity for tritium.…”

“…While their system was capable of imaging tritiated labeled gels, neither of their papers gave measurements of the efficiency or linearity for tritium. Burbeck et al [10] state a detector background noise of 0.95 cpm/mm 2 but did not give a value for their lower level discriminator. From noise curves for ªstandardº MCPs [8,12] we estimate they used a threshold of~0.6 times the peak detector gain.…”

Computerized analysis of two-dimensional electrophoretograms

“…When such numbers of proteins are separated, as they usually are in biological work, their quantification by automated procedures is almost a necessity. The first attempts at developing automated quantification procedures were published in 1978 (2)(3)(4), and several laboratories are now in various stages of developing their own such procedures (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20). The potential of a fully computerized system is outlined in ref 20, along with a short review of the field.…”