quantitative-pcr-of-t7-bacteriophage-from-biopanning

Quantitative PCR of T7 Bacteriophage from Biopanning

This protocol describes the use of quantitative PCR (qPCR) to enumerate T7 phages from phage selection experiments (i.e., &#34;biopanning&#34;). qPCR is a ...

University of Texas at Austin

Mucus is a selective barrier to particles and molecules, preventing penetration to the epithelial surface of mucosal tissues. Significant advances in transmucosal drug delivery have recently been made and have emphasized that an understanding of the basic structure, viscoelastic properties, and interactions of mucus is of great value in the design of efficient drug delivery systems. Mucins, the primary non-aqueous component of mucus, are polymers carrying a complex and heterogeneous structure with domains that undergo a variety of molecular interactions, such as hydrophilic/hydrophobic, hydrogen bonds and electrostatic interactions. These properties are directly relevant to the numerous mucin-associated diseases, as well as delivering drugs across the mucus barrier. Therefore, in this review we discuss regional differences in mucus composition, mucus physicochemical properties, such as pore size, viscoelasticity, pH, and ionic strength. These factors are also discussed with respect to changes in mucus properties as a function of disease state. Collectively, the review seeks to provide a state of the art roadmap for researchers who must contend with this critical barrier to drug delivery.

Physicochemical properties of mucus and their impact on transmucosal drug delivery.

Pseudomonas aeruginosa is the predominant pathogen in the persistent lung infections of cystic fibrosis (CF) patients among other diseases. One of the mechanisms of resistance of P. aeruginosa infections is the formation and presence of biofilms. Previously, we demonstrated that PEGylated-tobramycin (Tob-PEG) had superior antimicrobial activity against P. aeruginosa biofilms compared to tobramycin (Tob). The goal of this study was to optimize the method of PEGylation of Tob and assess its activity in an in vitro CF-like mucus barrier biofilm model. Tob was PEGylated using three separate chemical conjugation methods and analyzed by H NMR. A comparison of the Tob-PEG products from the different conjugation methods showed significant differences in the reduction of biofilm proliferation after 24 h of treatment. In the CF-like mucus barrier model, Tob-PEG was significantly better than Tob in reducing P. aeruginosa proliferation after only 5 h of treatment ( p < 0.01). Finally, Tob-PEG caused a reduction in the number of surviving P. aeruginosa biofilm colonies higher than that of Tob ( p < 0.0001). We demonstrate the significantly improved antimicrobial activity of Tob-PEG against P. aeruginosa biofilms compared to Tob using two PEGylation methods. Tob-PEG had better in vitro activity compared to that of Tob against P. aeruginosa biofilms growing in a CF-like mucus barrier model.

PEGylation of Tobramycin Improves Mucus Penetration and Antimicrobial Activity against Pseudomonas aeruginosa Biofilms in Vitro.

Peptides are small biological molecules that are attractive in drug delivery and materials engineering for applications including therapeutics, molecular building blocks and cell-targeting ligands. Peptides are small but can possess complexity and functionality as larger proteins. Due to their intrinsic properties, peptides are able to overcome the physiological and transport barriers presented by diseases. In this review, we discuss the progress of identifying and using peptides to shuttle across biological barriers and facilitate transport of drugs and drug delivery systems for improved therapy. Here, the focus of this review is on rationally designed, phage display peptides, and even endogenous peptides as carriers to penetrate biological barriers, specifically the blood-brain barrier(BBB), the gastrointestinal tract (GI), and the solid tumor microenvironment (T). We will discuss recent advances of peptides as drug carriers in these biological environments. From these findings, challenges and potential opportunities to iterate and improve peptide-based approaches will be discussed to translate their promise towards the clinic to deliver drugs for therapeutic efficacy.

Jasmim Leal

Division of Molecular Pharmaceutics and Drug Delivery,
College of Pharmacy,
Division of Molecular Pharmaceutics and Drug Delivery, College of Pharmacy

University of Texas at Austin

Quantitative PCR of T7 Bacteriophage from Biopanning

Jasmim Leal

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Division of Molecular Pharmaceutics and Drug Delivery,College of Pharmacy,Division of Molecular Pharmaceutics and Drug Delivery, College of Pharmacy

University of Texas at Austin

Quantitative PCR of T7 Bacteriophage from Biopanning

Division of Molecular Pharmaceutics and Drug Delivery,
College of Pharmacy,
Division of Molecular Pharmaceutics and Drug Delivery, College of Pharmacy