Abstract
Recently, the pump-probe technique has found highly valued developments and applications in optical microscopy to study both labeled (fluorescence) and non-labeled (vibrational) for observation of cells and tissues. In this work, we use the mechanism of stimulated emission (SE) with dual frequency modulation to remove the large background signal that is originated from spontaneous emission. We have applied a high performance digital signal processing based lock-in detection for stimulated emission fluorescence microscopy. Critically, the pump beam is modulated at f1 and the probe beam is at a different frequency, f2, with the demodulation carried out at the sum of the two frequencies, (f1 + f2). In this way, the DC background is effectively removed. Note that the background is mainly attributed to the spontaneous emission of the pump beam, with modulated frequency f1.