A commercially available computed radiography (CR) system for dental radiography was used to produce images from radiolabeled polyacrilamide gel electrophoresis (PAGE) assays. Typically, similar investigations require specific and expensive autoradiography devices. The CR unit was characterized in terms of sensitivity and fading by means of a 90Sr source that well simulates the experimental conditions, and then used for quantitative analyses of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) polymerase-independent ribonuclease H (RNase H) activity monitored by PAGE analysis. The results showed that the present methodology allows quantifying effectively the RNase H catalyses and that the obtained data are in good agreement with previous reference works. Finally, in order to further validate the present method in terms of relationship between enzyme activity, the rate of products formation and signal intensity, a PAGE analyses of the HIV-1 RNase H inhibition by the known diketo acid derivative RDS1643 was carried out.
Validation of a computed radiography device to monitor the HIV-1 RNase H activity
ESPOSITO, FRANCESCA;FANTI, VIVIANA;RANDACCIO, PAOLO;TRAMONTANO, ENZO;ZINZULA, LUCA
2009-01-01
Abstract
A commercially available computed radiography (CR) system for dental radiography was used to produce images from radiolabeled polyacrilamide gel electrophoresis (PAGE) assays. Typically, similar investigations require specific and expensive autoradiography devices. The CR unit was characterized in terms of sensitivity and fading by means of a 90Sr source that well simulates the experimental conditions, and then used for quantitative analyses of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) polymerase-independent ribonuclease H (RNase H) activity monitored by PAGE analysis. The results showed that the present methodology allows quantifying effectively the RNase H catalyses and that the obtained data are in good agreement with previous reference works. Finally, in order to further validate the present method in terms of relationship between enzyme activity, the rate of products formation and signal intensity, a PAGE analyses of the HIV-1 RNase H inhibition by the known diketo acid derivative RDS1643 was carried out.
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