Overcharging effect in electrospray ionization mass spectra of daunomycin-tuftsin bioconjugates

Peptide-based small molecule drug conjugates for targeted tumor therapy are currently in the focus of intensive research. Anthracyclines, like daunomycin, are commonly used anticancer drug molecules and are also often applied in peptide-drug conjugates. However, lability of the O-glycosidic bond during electrospray ionization mass spectrometric analysis hinders the analytical characterization of the constructs. “Overprotonation” can occur if daunomycin is linked to positively charged peptide carriers, like tuftsin derivatives. In these molecules, the high number of positive charges enhances the in-source fragmentation significantly, leading to complex mass spectra composed of mainly fragment ions. Therefore, we investigated different novel tuftsin-daunomycin conjugates to find an appropriate condition for mass spectrometric detection. Our results showed that shifting the charge states to lower charges helped to keep ions intact. In this way, a clear spectrum could be obtained containing intact protonated molecules only. Shifting of the protonation states to lower charges could be achieved with the use of appropriate neutral volatile buffers and with tuning the ion source parameters

3D printed microchannels for sub-nL NMR spectroscopy

Nuclear magnetic resonance (NMR) experiments on subnanoliter (sub-nL) volumes are hindered by the limited sensitivity of the detector and the difficulties in positioning and holding such small samples in proximity of the detector. In this work, we report on NMR experiments on liquid and biological entities immersed in liquids having volumes down to 100 pL. These measurements are enabled by the fabrication of high spatial resolution 3D printed microfluidic structures, specifically conceived to guide and confine sub-nL samples in the sub-nL most sensitive volume of a single-chip integrated NMR probe. The microfluidic structures are fabricated using a two-photon polymerization 3D printing technique having a resolution better than 1 \u3bcm3. The high spatial resolution 3D printing approach adopted here allows to rapidly fabricate complex microfluidic structures tailored to position, hold, and feed biological samples, with a design that maximizes the NMR signals amplitude and minimizes the static magnetic field inhomogeneities. The layer separating the sample from the microcoil, crucial to exploit the volume of maximum sensitivity of the detector, has a thickness of 10 \u3bcm. To demonstrate the potential of this approach, we report NMR experiments on sub-nL intact biological entities in liquid media, specifically ova of the tardigrade Richtersius coronifer and sections of Caenorhabditis elegans nematodes. We show a sensitivity of 2.5x1013spins/ Hz1/2on1H nuclei at 7 T, sufficient to detect 6 pmol of1H nuclei of endogenous compounds in active volumes down to 100 pL and in a measurement time of 3 hours. Spectral resolutions of 0.01 ppm in liquid samples and of 0.1 ppm in the investigated biological entities are also demonstrated. The obtained results may indicate a route for NMR studies at the single unit level of important biological entities having sub-nL volumes, such as living microscopic organisms and eggs of several mammalians, humans included

An exploratory study on strengthening and thermal stability of magnetron sputtered W nanoparticles at the interface of Cu/Ni multilayer films

An initial study to investigate the effect of controlled deposition of nanoparticles at multilayer interfaces was conducted to explore the mechanical effect of particles on laminate structures. Nanoparticles with diameter of about 4.5 nm were specifically deposited at the interface between Cu and Ni laminates by forced agglomeration of magnetron sputtered ions using a Mantis Ltd. Nanogen50 nanoparticle generator and the hardness of these films were measured using the nanoindentation technique. Cu/Ni laminates without W nanoparticles have an average modulus value of approximately 120 ± 3.7 GPa and hardness value of 2.23 ± 0.07 GPa, while the hardness values of the particle-containing films are greater, regardless of particle density. The areas with the lowest particle density at the interfaces (0.9 at.% W) show the greatest increase in hardness, with an increase of about 1.3 GPa greater than the particle-free sample. However, as the particle density increases, there is a corresponding decrease in hardness. In-situ x-ray diffraction of these films was also conducted to observe the annealing behavior of these films. For all samples, the Cu and Ni layered structure remained intact; however, there is evidence of Ni diffusion along grain boundaries and interaction with the oxygen, likely creating NiO. After annealing, a significant number of the W nanoparticles dissolved into the Ni matrix to create NiW solid-solution. The ability to deposit particles with such precise control has the potential to open up an exciting new field of research.publishedVersionPeer reviewe