In vitro aerosol characterization of Staccato(®) Loxapine.

Medicinal aerosol products (metered dose and dry powder inhalers) require characterization testing over a wide range of use and pre-operating stress scenarios in order to ensure robust product performance and support submissions for regulatory approval. Aerosol characterization experiments on Staccato(®) Loxapine for inhalation (Staccato Loxapine) product (emitted dose, particle size, and purity) were assessed at different operating settings (flow rates, ambient temperature and humidity, altitude, and orientation) and at nominal test conditions following exposure to various stresses on the device (mechanical shock, vibration, drop, thermal cycling, and light exposure). Emitted dose values were approximately 90% of the coated dose at every condition, meeting target specifications in each case. Aerosol purity was consistently >99.5% for every test setting, with no reportable impurities according to ICH standards (>0.1%). Particle size averaged 2µm (MMAD) and was independent of the different test conditions with the exception of different airflow rates. Particle size decreased slightly with airflow, which may assist in maintaining constant deep lung deposition. The combination of high emitted dose efficiency and a particle size range ideally suited for lung deposition, along with the consistency of these key aerosol attributes, suggests that the Staccato system has distinct advantages over more traditional aerosol systems.

Kinetics of oligonucleotide hybridization to DNA probe arrays on high-capacity porous silica substrates.

We have investigated the kinetics of DNA hybridization to oligonucleotide arrays on high-capacity porous silica films that were deposited by two techniques. Films created by spin coating pure colloidal silica suspensions onto a substrate had pores of approximately 23 nm, relatively low porosity (35%), and a surface area of 17 times flat glass (for a 0.3-microm film). In the second method, latex particles were codeposited with the silica by spin coating and then pyrolyzed, which resulted in larger pores (36 nm), higher porosity (65%), and higher surface area (26 times flat glass for a 0.3-microm film). As a result of these favorable properties, the templated silica hybridized more quickly and reached a higher adsorbed target density (11 vs. 8 times flat glass at 22 degrees C) than the pure silica. Adsorption of DNA onto the high-capacity films is controlled by traditional adsorption and desorption coefficients, as well as by morphology factors and transient binding interactions between the target and the probes. To describe these effects, we have developed a model based on the analogy to diffusion of a reactant in a porous catalyst. Adsorption values (k(a), k(d), and K) measured on planar arrays for the same probe/target system provide the parameters for the model and also provide an internally consistent comparison for the stability of the transient complexes. The interpretation of the model takes into account factors not previously considered for hybridization in three-dimensional films, including the potential effects of heterogeneous probe populations, partial probe/target complexes during diffusion, and non-1:1 binding structures. The transient complexes are much less stable than full duplexes (binding constants for full duplexes higher by three orders of magnitude or more), which may be a result of the unique probe density and distribution that is characteristic of the photolithographically patterned arrays. The behavior at 22 degrees C is described well by the predictive equations for morphology, whereas the behavior at 45 degrees C deviates from expectations and suggests that more complex phenomena may be occurring in that temperature regime.

Isoelectric focusing technology quantifies protein signaling in 25 cells.

A previously undescribed isoelectric focusing technology allows cell signaling to be quantitatively assessed in <25 cells. High-resolution capillary isoelectric focusing allows isoforms and individual phosphorylation forms to be resolved, often to baseline, in a 400-nl capillary. Key to the method is photochemical capture of the resolved protein forms. Once immobilized, the proteins can be probed with specific antibodies flowed through the capillary. Antibodies bound to their targets are detected by chemiluminescence. Because chemiluminescent substrates are flowed through the capillary during detection, localized substrate depletion is overcome, giving excellent linearity of response across several orders of magnitude. By analyzing pan-specific antibody signals from individual resolved forms of a protein, each of these can be quantified, without the problems associated with using multiple antibodies with different binding avidities to detect individual protein forms.

Kinetics of oligonucleotide hybridization to photolithographically patterned DNA arrays.

The hybridization kinetics of oligonucleotide targets to oligonucleotide probe arrays synthesized using photolithographic fabrication methods developed by Affymetrix have been measured. Values for the fundamental adsorption parameters, k(a), k(d), and K, were determined at both room temperature and 45 degrees C by monitoring the hybridization of fluorescently labeled targets to the array. The values for these parameters and the adsorbed target density (