A novel DNA platform designed for vaccine use with high transgene expression and immunogenicity.

The development of new, low-cost vaccines and effective gene therapies requires accurate delivery and high-level expression of candidate genes. We developed a plasmid vector, pIDV-II, that allows for both easy manipulation and high expression of exogenous genes in mammalian cells. This plasmid is based upon the pVax1 plasmid and shares a common structure with typical mammalian transcription units. It is composed of a chicken ß-actin promoter (CAG), followed by an intron and flanked by two restriction sites, and also includes a post-transcriptional regulatory element, followed by a transcriptional termination signal. While the modification of pVax1 elements either decreased eGFP expression levels or had no effect at all, replacement of the promoter, the poly-A signal, deletion of the T7 and AmpR promoters, and inversion of the ORI-Neo/Kan cassette, significantly increased in vitro eGFP expression with the modified plasmid called pIDV-II. To further evaluate our vector, expression levels of three viral antigens were compared in cell lines transfected either with pVax1 or pCAGGS backbones as controls. Higher transgene expression was consistently observed with pIDV-II. The humoral and cellular responses generated in mice immunized with pIDV-II vs pVax1 expressing each viral antigen individually were superior by 2-fold or more as measured by ELISA and ELISPOT assays. Overall these results indicate that pIDV-II induces robust transgene expression, with concomitant improved cellular and humoral immune responses against the transgene of interest over pVax1. The new vector, pIDV-II, offers an additional alternative for DNA based vaccination and gene therapy for animal and human use.

Surgical outcomes of endothelial keratoplasty safety-basket suture for management of difficult cases.

OBJECTIVE: To report outcomes of Descemet stripping automated endothelial keratoplasty (DSAEK) safety-basket suture technique in complex eyes in which an air bubble could not be maintained in the anterior chamber. STUDY DESIGN: Retrospective chart review. PARTICIPANTS: Patient's undergoing DSAEK with use of the safety-basket suture technique in patients with corneal edema secondary to endothelial dysfunction with up to 3 years of follow-up between 2013 and 2016. METHODS: Outcome measures were corrected distance visual acuity, average endothelial cell count loss, graft decentration, graft detachment, rebubble rate, infection rate, graft rejection, and graft failure were analyzed. RESULTS: The technique was used successfully for 36 high-risk cases with no incidence of graft decentration, postoperative infection, or graft rejection. Both the graft detachment and rebubble rate was found to be 11.1%. The graft failure rate was found to be 22.2% with an average of 3.3 months, with 37.5% of these being primary failure and 62.5% of these being secondary failure. CONCLUSIONS: Endothelial keratoplasty safety-basket suture is a technique that can be beneficial in difficult DSAEK cases with a lower graft detachment rate compared with the current literature as well as a fairly low complication rate and graft failure while maintaining good visual outcomes. The safety-basket suture is the technique of choice for cases of aphakia and may allow for DSAEK in cases previously that may have only been considered for full-thickness keratoplasty.

Endothelial Keratoplasty Safety-Basket Suture for Management of Difficult Cases.

PURPOSE: To describe a novel technique of a safety-basket for management of Descemet stripping automated endothelial keratoplasty in eyes with defects of the lens-iris diaphragm, postvitrectomized eyes, eyes with previous glaucoma tube-shunt or trabeculectomy, or when the patient cannot lie supine. METHODS: The safety-basket suture is placed after the endothelial graft has been positioned under air and all wounds have been closed. Double-armed 10-0 polypropylene suture on a curved needle is passed under the graft in a single-running fashion, a modification of the Masket technique used as a safety suture in the management of malpositioned intraocular lenses. Care is taken not to touch the graft with the needle or suture. The suture can be left in place and removed at the slit-lamp postoperatively with continued use of topical antibiotics while the safety-basket is left in place. RESULTS: The technique was used successfully for 35 high-risk cases with good success with no cases of postoperative infection. CONCLUSIONS: Endothelial keratoplasty safety-basket sutures, when used in high-risk Descemet stripping automated endothelial keratoplasty patients with abnormal anterior segment anatomy can provide support to maintain the graft in a central position and thus allow for easier rebubbling, should it detach in the early postoperative course.

Pyrene-functionalised, alternating copolyimide for sensing nitroaromatic compounds.

A novel, pyrene-functionalised copolymer has been synthesised in a single step via imidisation of poly(maleic anhydride-alt-1-octadecene) with 1-pyrenemethylamine, and its potential for the detection of volatile nitro aromatic compounds (NACs) evaluated. The new copolymer forms complexes in solution with NACs such as 2,5-dinitrobenzonitrile, as shown by (1) H NMR, UV-vis and fluorescence spectroscopy. Moreover, thin films of this copolymer, cast from THF solution, undergo almost instantaneous fluorescence quenching when exposed to the vapour of 2,5-dinitrobenzonitrile (a model for TNT) at ambient temperatures and pressures.