Observing protein synthesis, export, and tryptophan incorporation by front-surface fluorescence.
Front-surface detection of emission from fluorophores in the presence and absence of light-scattering particles was contrasted to right-angle and wave-guide detection. We found that front-surface detection was the least prone to the reabsorption, inner-filtering, and scattering effects that can plague fluorescent measurements. Front-surface detection was thus used to assess the use of protein and ANS fluorescence as a means of monitoring events in bacterial fermentations. Protein fluorescence appeared to track well changes in optical density during balanced growth. However, during the lag associated with diauxic growth and after exposure to ampicillin, protein fluorescence became decoupled from cellular growth in a manner consistent with prior observations and the known effect of ampicillin on cells. ANS proved to be nontoxic and capable of reporting the occurrence of protein release from cells. The spectral shifts of tryptophan indicated that the incorporation of tryptophan into cellular protein can be monitored.
Lipton
A J
AJ
Department of Chemical Engineering, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213, USA.
Domach
M M
MM