Nanoform of Phospholipid Composition: Investigation of the Morphological Features by Atomic Force Microscopy.

Morphological features of the nanoform of a phospholipid composition (NFPh), which can be used as an individual pharmaceutic agent or as a platform for designing drug delivery systems, have been studied using atomic force microscopy (AFM). NFPh has been developed, and its characteristics have been investigated using conventional drug analysis methods, including the determination of the mean diameter of nanosized vesicles in the emulsion via dynamic light scattering (DLS). Using DLS, the mean diameter of the vesicles was found to be ~20 nm. AFM imaging of the surface has revealed four types of objects related to NFPh: (1) compact objects; (2) layer fragments; (3) lamellar structures; and (4) combined objects containing the compact and extended parts. For type (4) objects, it has been found that the geometric ratio of the volume of the convex part to the total area of the entire object is constant. It has been proposed that these objects formed owing to fusion of vesicles of the same size (with the same surface-to-volume ratio). It has been shown that this is possible for vesicles with diameters of 20 nm. This diameter is in good coincidence with the value obtained using DLS.

Plasma preparation to measure FDA-approved protein markers by selected reaction monitoring.

BACKGROUND: The development of commercially available panels for human blood plasma screening via selected reaction monitoring (SRM) offers reliable, cost-efficient and highly-standardized discovery and validation of protein biomarkers. However, protein detection by SRM can be hampered by interfering peptide fragment ions. To estimate the influence of interference on protein detection, we performed different types of sample preparation and implemented SRM measurements for well-characterized protein targets approved by the US Food and Drug Administration. METHODS: We used the PlasmaDeepDive™ SRM assay from BiognoSYS AG for absolute quantification of 18 proteins in 19 samples of human plasma using three different protocols for sample preparation. SRM measurements were performed using iRT standards for retention time normalization and isotopically-labeled reference peptides for absolute quantification. SpectroDive™ software was used for automated detection of reliable peak groups. RESULTS: Fourteen targeted proteins were quantitatively measured in more than half of the samples. Depletion of highly-abundant plasma proteins and peptide fraction clean-up on centrifuge plates resulted in detection of all 18 targeted proteins in femtomolar to picomolar concentrations. CONCLUSIONS: It was shown that commercially designed SRM kits are suitable for SRM detection of well-established plasma/serum biomarkers.