Transformation of seven species of filamentous fungi using the nitrate reductase gene of Aspergillus nidulans

A gene transfer system originally developed for Fusarium oxysporum has been applied to seven species of filamentous fungi of agricultural and industrial importance. This transformation system relies on the selection of mutants deficient in nitrate reductase by positive screening. Such mutants were recovered easily in all the fungi tested--without mutagenic treatments--through their resistance to chlorate. They were transformed by a plasmid vector (pAN301) carrying the Aspergillus nidulans wild-type gene (niaD). Transformation frequencies ranged from one to ten transformants/micrograms plasmid DNA. The general properties of the transformants were analyzed. Most of them are mitotically stable, and the integration of the vector into the host genome frequently occurred in a tandem fashion.     

Expandable intraluminal graft: a preliminary study. Work in progress

To overcome the problem of recurrence of stenosis after vascular balloon dilatations, we developed an expandable, intraluminal graft that allows dilatation of the lesion and simultaneous placement of a supportive endoprosthesis to prevent recoil of the arterial wall. The graft is made of continuous, woven, stainless steel wire. The resulting tubular mesh has a wall thickness of 200-450 micron and 80% open surface. The grafts, mounted on angioplasty catheters, are introduced through 8-12-F Teflon sheaths. Eleven grafts of 6, 8, and 10 mm in diameter by 20 mm long were placed in the aorta, common carotid, superior mesenteric, iliac, and renal arteries of dogs. Six grafts showed no stenosis in follow-up studies of up to 8 weeks. Two grafts had moderate stenosis as a result of neointimal hyperplasia. Two partial and one complete graft thrombosis occurred in nonheparinized animals in which the graft outflow was restricted. Anticoagulant was not used on a long-term basis. Light and electron microscopy studies showed complete covering of the graft's inner surface by endothelium at 3 weeks.     

Quantitation of residual white cells in filtered blood components by polymerase chain reaction amplification of HLA DQ-A DNA

BACKGROUND: Over the past several years, blood filtration technology has improved dramatically, such that currently available experimental filters are capable of reducing white cells (WBCs) in blood components to less than 0.1 WBC per microL. These residual WBC concentrations are below the sensitivity of automated cell counters, as well as of large- volume (Nageotte) hemocytometers. STUDY DESIGN AND METHODS: A quantitative polymerase chain reaction (PCR) amplification assay directed at HLA DQ-A DNA sequences has been developed for the enumeration of WBCs in filtered blood. To ensure quantitative recovery of WBCs at very low residual cell concentrations, a direct red cell lysis and WBC concentration protocol using 0.5 mL of filtered blood was perfected. Amplified product is detected by oligomer hybridization using 32P-labeled probes, with quantitation by image analysis of autoradiographic signals relative to a standardized dilution series processed in parallel. RESULTS: Recovery of residual WBCs in filtrates was shown to be enhanced by the addition of xenogeneic WBCs or polystyrene beads, which served as "carrier" particles during red cell lysis and wash steps. A contribution of nuclear fragments in filtered blood to PCR signal in the range of 0.01 to 0.5 WBCs per microL was observed; a modified protocol was developed to minimize this effect. Parallel analysis of spiked dilution series and evaluations of 39 red cell components filtered through commercial filters indicated good correlation between PCR and standard Nageotte counts in the range of 0.1 to 10 WBCs per microL (r2 = 0.94); only PCR was able to detect residual WBCs in filtrates from prototype 6 log10 WBC-reduction filters. CONCLUSION: This assay should prove useful for the development and quality assurance of increasingly efficient WBC-reduction filters.     

A latex particle assay for platelet-associated IgG

A modified, sensitive, solid-phase assay for platelet-associated IgG is reported. Direct comparisons were made between a 125I monoclonal radioimmunoassay (RIA) and the polyclonal antibody latex particle assay. In 209 simultaneous comparisons with the RIA, the sensitivity of the latex test was 91 percent, specificity was 100 percent, and overall efficiency was 97 percent. Commencing with platelet-rich plasma, the direct latex particle test for platelet-bound IgG requires 45 minutes; 90 minutes are required to crossmatch one patient with 12 donors. The advantages of the latex assay are absence of radioactivity, stability of reagents, economy, speed, specificity, and sensitivity.