ONLINE GREEN FLUORESCENT PROTEIN SENSOR WITH LED EXCITATION

We present an intensity based sensor designed for on-line monitoring o f green fluorescent protein, a revolutionary marker of protein express ion. The device consisted of a blue light emitting diode as the excita tion source. A band pass excitation filter cut off light longer than 4 90 nm. The light was directed into a bifurcated optical fiber bundle w ith the common end inserted into a stainless steel housing equipped wi th a quartz window. The fiber bundle and stainless steel housing are s team sterilizable. The emission radiation was collected through a long wave pass filter to reject the excitation light shorter than 505 nm a nd was detected by a photomultiplier tube. The signal was amplified an d sent to a computer for recording time course data. The sensor was te sted in an Escherichia coli fermentation of JM105 transformed with pBA D-GFP. The on-line signal was compared to off-line fluorescence spectr ophotometer measurements. The on-line profile closely followed the off -line. Western blot data showed that with a time shift, the sensor was able to both continuously and quantitatively monitor expression of gr een fluorescent protein on-line in real time. (C) 1997 John Wiley & So ns, Inc.