Development of an advanced multiwavelength emission detector for the analytical ultracentrifuge

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Development of an advanced multiwavelength emission detector for the analytical ultracentrifuge.

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Abstract:

An advanced design of the analytical ultracentrifuge (AUC) with multiwavelength emission detection (MWE-AUC) is presented which offers outstanding performance concerning the spectral resolution and range flexibility as well as the quality of the data acquired. The excitation by a 520 nm laser is complemented with a 405 nm laser. An external spectrograph with three switchable tunable gratings permits optimisation of the spectral resolution in an order of magnitude range while keeping the spectral region broad. The new system design leads also to a significant reduction of systematic signal noise and allows the assessment and control of inner filter effects. Details regarding the very large signal dynamic range are presented, an important aspect when studying samples in a broad concentration range of up to five orders of magnitude. Custom-made mono-sector sample cells with 3 mm titanium centerpiece have been developed and are applied as default system components. Our system is validated by complementary studies on two biological systems, fluorescent bovine serum albumin and green fluorescent protein, using the commercial Optima AUC with absorbance detection for comparison. Finally, we demonstrate the capabilities of our second generation MWE-AUC with respect to multiwavelength characterisation of gold nanoclusters, which exhibit specific fluorescence depending on their structure. Overall, this work depicts an important step-stone for the future in-depth studies of size-, shape- and composition-dependent multiwavelength emission properties of colloids.