The entire surface protein of hepatitis B virus serotype ayw containing the preS (preS1+preS2) and S domains has been expressed in the yeast Saccharomyces cerevisiae. Yeast containing a recombinant plasmid utilizing a constitutive promoter did not express this gene successfully due to the toxicity of the protein. A plasmid using a regulatable promoter directed expression which initiated late in the exponential phase of growth and resulted in the accumulation of high intracellular levels of the complete surface protein. The purified polypeptide aggregates into a form which, although not comprised of typical 20 nm particles, displays antigenic determinants encoded by the preS1, preS2 and S domains. Immunization of rabbits elicited the formation of antibodies directed against all three domains. This candidate vaccine will be useful for studying the contributions to viral immunity of the host response to the preS1 and preS2 domains.

Animals↗

PubMedDate not supplied

The uses of computer-aided signal peptide selection and polymerase chain reaction in gene construction and expression of secreted proteins.

The computer program, SIGSEQ2, was used to select heterologous signal peptides from a catalog of published sequences to express the echistatin gene in insect (Sf9) cells. S-values for each amino acid were determined to select empirically the site of cleavage between the signal peptide and mature echistatin. Five gene fragments encoding the signal peptides for human immunoglobulin kappa (Ig kappa), Drosophila 68C glue, antistasin, bovine growth hormone (bGH), and human apolipoprotein E (Apo E) were constructed by the use of long synthetic oligonucleotides or polymerase chain reaction (PCR). Echistatin expression vectors then were constructed using the baculovirus polyhedrin promoter. Following transient transfection, the media were assayed for echistatin activity. The results indicate that the computer program greatly facilitated the selection and design of five different signal peptides and accurately predicted their relative functionality in the expression and secretion of echistatin in insect cell cultures.

Animals↗

PubMedDate not supplied

Effect of air abrasion and acid etching on sealant retention: an in vitro study.

PURPOSE: This in-vitro study evaluated shear bond strength and analyzed, via scanning electron microscopy, enamel prepared for pit and fissure sealant placement. Various surface pretreatment methods were conducted prior to short-term (72-hour) and long-term (120-day) analysis. METHOD: Six treatment groups of 14 teeth, having 28 bonding surfaces (buccal and lingual) were treated. Cylinders of Delton pit and fissure sealant were placed on the prepared buccal and lingual surfaces and light-cured for 60 seconds. One-half of each group's specimens were stored in distilled water for 72 hours and the other half were stored in water distilled for 120 days followed by thermocycling. All specimens were subjected to shear bond strength analysis as determined on an Instron testing machine. RESULTS: All acid treated groups were equivalent and greater than air abrasion alone after 72 hours of water storage. Scanning electron microscopy of air-abraded and combination treated enamel surfaces revealed a more detailed retentive pattern in the combination treatment than in either treatment alone. CONCLUSION: Based on in-vitro shear bond strength values, air abrasion with 50 micron alumina is an effective pre-etch treatment for sealant placement and in concert with phosphoric acid treatment significantly enhanced the long-term bond of a sealant to enamel. The clinical relevance of this has not been established.

Acid Etching, Dental↗

PubMedDate not supplied

New vaccine technologies.

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Animals↗