Prometheus Panta is equipped with an advanced optical module which enables recording of dynamic light scattering (DLS) data. DLS measurements can be conducted isothermally to determine particle size, sample homogeneity, and weak particle interactions, or simultaneous to nanoDSF and turbidity measurements during a thermal ramp in a melt scan to determine how the thermal stress affects the particle size and homogeneity of the sample. These independent and equally important measurements can be performed on the same sample, with the capacity to measure up to 48 samples in a single experiment.

The DLS module is equipped with an advanced componentry, including a cutting-edge laser and detector, to enable light scattering measurements to be conducted in parallel to nanoDSF and backreflection measurements with no compromise to the quality of results or thermal control. The DLS laser in Prometheus Panta illuminates the samples at 405 nm. The wavelength of 405 nm results in an increased scattering intensity, enabling the DLS measurement to record high quality results while using a smaller scattering volume and shorter acquisition time. Please note that samples containing molecules that absorb or fluoresce in the range of the laser wavelength (e.g. GFP) are not compatible with DLS measurements on Prometheus Panta.

The scattering volume, the part of the sample where the DLS optic is focused to measure the fluctuations in scattering intensity caused by Brownian motion of the particles in solution (see illustration below), is focused just beneath the capillary wall. This small volume (60 x 30 x 30 μm) is readjusted for every DLS acquisition using the fluorescence signal from the capillary to eliminate artefacts caused by reflections or scattering from the capillary wall. In addition to enabling high quality DLS measurements in the smaller inner-diameter capillaries used on Prometheus Panta with optimal thermal transfer properties, use of a small scattering volume also decreases the effect of multiple scattering events from concentrated samples, but does increase the susceptibility of the measurement to the effect of number fluctuations.

The scattered light is detected with a modern photomultiplier tube (PMT) that is instrumental to the fast acquisition of high-quality results. The PMT in the optical module is positioned in a backscatter orientation (see illustration below). As larger particles have increased scattering cross sections in the forward direction according to Mie theory, the orientation of the PMT increases the sensitivity of the system to small biologic samples (including proteins, antibodies, and viruses), and decreases the effect of dust and aggregates. Additionally, the PMT in the optical module can dynamically switch between photon counting and analog sampling modes to enable seamless measurement of both weakly and strongly scattering samples. While all the photons need to be counted to precisely characterize weakly scattering samples, as samples aggregate and their scattering intensity increases, they can very rapidly saturate detectors that operate in photon counting mode. By dynamically switching to an analogue mode, the DLS optical module in Prometheus Panta can continuously measure samples with varying scattering intensities. This enables measurement of particle size changes caused by events as diverse as the subtle increase in protein size upon unfolding to the drastic increase in particle size through aggregate formation, all while continuously heating up to 48 samples simultaneously.