Using enhanced development tools offered by analytical Quality by Design to support switching of a quality control method.

Quality by Design (QbD) principles play an increasingly important role in the pharmaceutical industry. Here, we used an analytical QbD (AQbD) approach to develop a capillary electrophoresis sodium dodecyl sulfate under reducing conditions (rCE-SDS), with the aim of replacing SDS-polyacrylamide gel electrophoresis (SDS-PAGE) as release and stability test method for a commercialized monoclonal antibody product. Method development started with defining analytical method performance requirements as part of an analytical target profile, followed by a systematic risk assessment of method input parameters and their relation to defined method outputs.

Real time in-line monitoring of large scale Bacillus fermentations with near-infrared spectroscopy.

NIR spectroscopy was used to monitor Bacillus fermentations in 50 m(3) reactors under harsh industrial conditions. The BioPAT(®) Spectro NIR sensor was attached directly to the bioreactor and provided fast, sensitive, non-destructive and robust measurements without interfering with the microorganism metabolism. Multivariate data analysis techniques related the spectra collected in real time during the fermentation with reference analyte concentrations.

Blinking effect and the use of quantum dots in single molecule spectroscopy.

Luminescent semiconductor nanocrystals (quantum dots, QD) have unique photo-physical properties: high photostability, brightness and narrow size-tunable fluorescence spectra. Due to their unique properties, QD-based single molecule studies have become increasingly more popular during the last years. However QDs show a strong blinking effect (random and intermittent light emission), which may limit their use in single molecule fluorescence studies.

Time-dependent FRET with single enzymes: domain motions and catalysis in H(+)-ATP synthases.

H(+)-ATP synthases are molecular machines which couple transmembrane proton transport with ATP synthesis from ADP and inorganic phosphate by a rotational mechanism. Single-pair fluorescence resonance energy transfer (spFRET) in single molecules is a powerful tool to analyse conformational changes. It is used to investigate subunit movements in H(+)-ATP synthases from E.

Subunit movements in single membrane-bound H+-ATP synthases from chloroplasts during ATP synthesis.

Subunit movements within the H(+)-ATP synthase from chloroplasts (CF(0)F(1)) are investigated during ATP synthesis. The gamma-subunit (gammaCys-322) is covalently labeled with a fluorescence donor (ATTO532). A fluorescence acceptor (adenosine 5'-(beta,gamma-imino)triphosphate (AMPPNP)-ATTO665) is noncovalently bound to a noncatalytic site at one alpha-subunit.

Blue luminescence and superstructures from magic size clusters of CdSe.

In this letter, we present a low-temperature synthesis route revealing a new type of ultrasmall CdSe nanoparticle family with exceptional narrow blue emissions between 437 and 456 nm and full width at half-maxima below 20 nm. Transmission electron microscopy characterization shows the uniformity of the nanoparticles, which have a diameter of 1.6 nm.

Fluorescence resonance energy transfer in single enzyme molecules with a quantum dot as donor.

H(+)-ATPsynthases couple a transmembrane proton transport with ATP synthesis and ATP hydrolysis. Previously, the relative subunit movement during this process has been measured by fluorescence resonance energy transfer (FRET) between two organic fluorophores covalently bound to different subunits. To improve the photophysical stability, a luminescent CdSe/ZnS nanocrystal (quantum dot) was bound to the enzyme and an organic fluorophore, Alexa568, was used as fluorescence acceptor.