Infrared spectroscopy is a versatile method for studying biological samples in liquid media. This led to numerous applications such as label-free identification of cells and bacteria, quantification of clinically relevant parameters in body fluids and early cancer detection using liquid biopsies.
Most of the studies performed with broadband spectroscopy have been performed with Fourier-Transform Infrared Spectrometers (FTIR), which are equipped with thermal light sources. However, the moderate power of the thermal light source and the low sensitivity of available mid-infrared detectors limit the achievable sensitivity. This problem is further aggravated for liquid or aqueous samples. Water is a strong absorber in the mid-infrared (MIR) wavelength range. Therefore, in the majority of cases the thickness of the sample is usually limited to ≈10 µm. Besides limiting the applicability to very thin samples, practical problems arise, like clogging of liquid cuvettes.
In a recent paper, our team has demonstrated in theory and experiment the potential of field-resolved infrared spectroscopy (FRS) to overcome these long-standing limitations. FRS relies on the excitation of resonant molecular vibrations with waveform-stable, broadband MIR pulses, and electric-field-resolved detection of the emerging fingerprint waveforms. FRS makes it possible to record a molecular signal with high signal-to-noise ratio, although water attenuates the excitation by orders of magnitude. We found that sub-µg/ml detection sensitivities can be maintained for samples as thick as 80 µm and that even for 0.2-mm-thick samples, detection sensitivities in the range of 10 µg/ml are feasible, which is the level achieved by state-of-the-art research-grade FTIR instruments, albeit under the stringent condition of sub-10-µm sample thickness.
The relaxed requirements to the sample thickness enable a more flexible design of liquid cuvettes and microfluidic chips for MIR spectroscopic applications. In addition, the increased signal-to-noise ratio for the measurement of thick aqueous samples will benefit MIR transmission spectroscopy and spectro-microscopy of biological samples, such as living cells, bulk cell and tissue-cultures as well as biological tissues, in their natural (hydrated) state.
Most of the studies performed with broadband spectroscopy have been performed with Fourier-Transform Infrared Spectrometers (FTIR), which are equipped with thermal light sources. However, the moderate power of the thermal light source and the low sensitivity of available mid-infrared detectors limit the achievable sensitivity. This problem is further aggravated for liquid or aqueous samples. Water is a strong absorber in the mid-infrared (MIR) wavelength range. Therefore, in the majority of cases the thickness of the sample is usually limited to ≈10 µm. Besides limiting the applicability to very thin samples, practical problems arise, like clogging of liquid cuvettes.
In a recent paper, our team has demonstrated in theory and experiment the potential of field-resolved infrared spectroscopy (FRS) to overcome these long-standing limitations. FRS relies on the excitation of resonant molecular vibrations with waveform-stable, broadband MIR pulses, and electric-field-resolved detection of the emerging fingerprint waveforms. FRS makes it possible to record a molecular signal with high signal-to-noise ratio, although water attenuates the excitation by orders of magnitude. We found that sub-µg/ml detection sensitivities can be maintained for samples as thick as 80 µm and that even for 0.2-mm-thick samples, detection sensitivities in the range of 10 µg/ml are feasible, which is the level achieved by state-of-the-art research-grade FTIR instruments, albeit under the stringent condition of sub-10-µm sample thickness.
The relaxed requirements to the sample thickness enable a more flexible design of liquid cuvettes and microfluidic chips for MIR spectroscopic applications. In addition, the increased signal-to-noise ratio for the measurement of thick aqueous samples will benefit MIR transmission spectroscopy and spectro-microscopy of biological samples, such as living cells, bulk cell and tissue-cultures as well as biological tissues, in their natural (hydrated) state.
