Flow imaging microscopy for protein particle analysis--a comparative evaluation of four different analytical instruments.

Flow imaging microscopy was introduced as a technique for protein particle analysis a few years ago and has strongly gained in importance ever since. The aim of the present study was a comparative evaluation of four of the most relevant flow imaging microscopy systems for biopharmaceuticals on the market: Micro-Flow Imaging (MFI)4100, MFI5200, Flow Cytometer And Microscope (FlowCAM) VS1, and FlowCAM PV. Polystyrene standards, particles generated from therapeutic monoclonal antibodies, and silicone oil droplets were analyzed by all systems.

Micro-flow imaging and resonant mass measurement (Archimedes)--complementary methods to quantitatively differentiate protein particles and silicone oil droplets.

Our study aimed to comparatively evaluate Micro-Flow Imaging (MFI) and the recently introduced technique of resonant mass measurement (Archimedes, RMM) as orthogonal methods for the quantitative differentiation of silicone oil droplets and protein particles. This distinction in the submicron and micron size range is highly relevant for the development of biopharmaceuticals, in particular for products in prefilled syringes. Samples of artificially generated silicone oil droplets and protein particles were quantified individually and in defined mixtures to assess the performance of the two techniques.

How subvisible particles become invisible-relevance of the refractive index for protein particle analysis.

The aim of the present study was to quantitatively assess the relevance of transparency and refractive index (RI) on protein particle analysis by the light-based techniques light obscuration (LO) and Micro-Flow Imaging (MFI). A novel method for determining the RI of protein particles was developed and provided an RI of 1.41 for protein particles from two different proteins.

Pharmaceutical feasibility of sub-visible particle analysis in parenterals with reduced volume light obscuration methods.

The draft for a new United States Pharmacopoeia (USP) monograph {787} "Sub-visible Particulate Matter in Therapeutic Protein Injections" describes the analysis of sub-visible particles by light obscuration at much lower sample volumes as so far required by the European Pharmacopoeia (Ph. Eur.) and the USP for parenterals in general.

Non-ionic Gd-based MRI contrast agents are optimal for encapsulation into phosphatidyldiglycerol-based thermosensitive liposomes.

Thermosensitive liposomes (TSL) with encapsulated magnetic resonance imaging (MRI) longitudinal relaxation time (T(1)) contrast agents (CAs) have been proposed for MRI assisted interventional thermotherapy in solid tumors. Here the feasibility of 6 clinically approved CAs (Gd-DTPA, Gd-BOPTA, Gd-DOTA, Gd-BT-DO3A, Gd-DTPA-BMA, and Gd-HP-DO3A) for formulation into TSL was investigated. CAs were passively encapsulated with 323 mOs kg(-1) into 1,2-dipalmitoyl-sn-glycero-3-phosphocholine/1,2-distearoyl-sn-glycero-3-phosphocholine/1,2-dipalmitoyl-sn-glycero-3-phosphodiglycerol 50/20/30 (mol/mol) TSL (DPPG(2)-TSL) to obtain stable formulations.

Implementation and evaluation of an optical fiber system as novel process monitoring tool during lyophilization.

Lyophilization is an important and well-established pharmaceutical drying process. Product temperature is the most critical process parameter during lyophilization as it impacts both product quality and process efficiency. Traditionally, thermocouples (TCs) or resistance temperature detectors (RTDs) and recently, manometric temperatures measurements (MTMs) have been used to monitor the product temperature.

Particles in therapeutic protein formulations, Part 1: overview of analytical methods.

The presence of particles is a major issue during therapeutic protein formulation development. Both proteinaceous and nonproteinaceous particles need to be analyzed not only due to the requirements of the Pharmacopeias but also to monitor the stability of the protein formulation. Increasing concerns about the immunogenic potential together with new developments in particle analysis make a comparative description of established and novel analytical methods useful.

Forced degradation of therapeutic proteins.

The scope of this paper is to review approaches used for forced degradation (synonym, stress testing) of therapeutic proteins. Forced degradation studies play a central role in the development of therapeutic proteins, for example, for candidate selection, molecule characterization, formulation development, assay development, and comparability studies. Typical stress methods are addressed within this review, such as exposure to elevated temperatures, freeze-thawing, mechanical stress, oxidation, light, as well as various materials and devices used in the clinics during final administration.

Size of thermosensitive liposomes influences content release.

Thermosensitive liposomes (TSL) in combination with regional hyperthermia represent a powerful tool for tumor specific drug delivery. The objective of this study was to investigate the influence of vesicle size on the biophysical properties of TSL. TSL were composed of DPPC/DSPC/1,2-dipalmitoyl-sn-glycero-3-phosphoglyceroglycerol (DPPG(2)) 50:20:30 (mol/mol) (DPPG(2)-TSL) and DPPC/P-Lyso-PC/DSPE-PEG2000 90:10:4 (mol/mol) (PEG/Lyso-TSL) with encapsulated fluorescent dye carboxyfluorescein, anticancer drug doxorubicin or magnetic resonance contrast agent gadodiamide.

In vitro stability and content release properties of phosphatidylglyceroglycerol containing thermosensitive liposomes.

Recently, we reported that 1,2-dipalmitoyl-sn-glycero-3-phosphoglyceroglycerol (DPPGOG) prolongs the circulation time of thermosensitive liposomes (TSL). Since the only TSL formulation in clinical trials applies DSPE-PEG2000 and lysophosphatidylcholine (P-lyso-PC), the objective of this study was to compare the influence of these lipids with DPPGOG on in vitro stability and heat-induced drug release properties of TSL. The content release rate was significantly increased by incorporating DPPGOG or P-lyso-PC in TSL formulations.