Performance and use of a ribonucleotide reductase herpes simplex virus type-specific serological assay.

In response to the increasingly evident need for herpes simplex virus (HSV) serotype-specific serologic assays that rely on proteins other than glycoprotein-G (gG), we developed a rapid serologic assay that is based on type-specific epitopes within the large subunit of HSV ribonucleotide reductase (R1). The assay (Au-2 enzyme-linked immunosorbent assay [ELISA]) uses an HSV type 2 (HSV-2) R1 peptide antigen. It provides a reliable method for detecting serotype-specific antibody to a protein other than gG-2. The Au-2 ELISA has high sensitivity and specificity as determined by direct comparison to Western blotting, a widely accepted "gold standard," and to ELISA with an HSV-1 R1 peptide (Au-1). The use of the Au-2 ELISA in conjunction with the gG-2-based assays will improve the sensitivity and specificity of serologic diagnosis and patient management.

Immobilization of enzymes with polyaziridines.

A novel method of enzyme immobilization using a low molecular weight prepolymer of tri-functional aziridines which can immobilize enzymes both by covalent attachment and entrapment within a gel matrix is described. The enzymes are immobilized on a solid support and exhibit an excellent retention of enzymatic activity. The immobilization procedure is essentially a single step process which can be easily performed at room temperature or 4 degrees C in either aqueous solution or in an inert organic solvent. The polyaziridines used in the immobilization are nontoxic, available in bulk at low cost and completely miscible with water and many organic solvents, thus providing one of the most satisfactory methods of immobilization available.

Separation of three exotoxic factors of Bacillus anthracis by sequential immunosorbent chromatography.

Protective antigen and lethal factor components were isolated directly from crude culture supernatant of Bacillus anthracis by sequential immunosorbent chromatography using immobilized monoclonal antibodies (MAB) against the respective toxins. The immunological activity of protective antigen, lethal factor and edema factor were purified by 1.2-, 6.3- and 2.3-fold, respectively, with recoveries of 63, 70 and 46%, respectively. All three components retained biological activity when combined to form lethal toxin or edema toxin, PA + LF and PA + EF, respectively, after the purification process, and were not contaminated with any of the other components. The order of immunosorbent columns during the purification process was found to be important. The best results were obtained when the protective antigen was removed initially from the crude culture supernatant.

Purification of Chironex fleckeri venom components using Chironex immunoaffinity chromatography.

A comparison of the purification of the nematocyst venom of Chironex fleckeri by affinity immunochromatography using 13 different monoclonal antibodies was made. Varying degrees of purification of mouse lethal factor, hemolysin and dermonecrotic factors, as well as antigen positive proteins were achieved with each of the monoclonal antibodies. Although the protein curves of the chromatography were similar, each of the monoclonal antibody columns had a distinctive pharmacological and SDS-PAGE profile. At least two hemolysins (120,000 and 70,000 molecular weight), two dermonecrotic principles (120,000, less than 120,000) and three lethal factors (120,000, 70,000 and 14,500 molecular weight) were detected. The degree to which aggregation and fragmentation affects the molecular weights of these proteins is not known. It appears that multiple pharmacological activities are present within the same molecule since it is only with great difficulty that a pharmacological activity can be assigned to a specific molecular weight.

Serological diagnosis of jellyfish envenomations.

1. A good correlation between the clinical and serological identification of envenomating jellyfish could be made on 30 healthy individuals and 74 patients stung by known species. 2. Six patients and one previous case were known to be false positive reactors. 3. Two of these individuals had dermatitis, one was asthmatic, three had previous significant hymenoptera envenomations and one was apparently normal. 4. Specific anti-jellyfish IgG serum concentrations would appear a few days after envenomation and persist for many months, even at high concentrations. 5. Significant numbers of patients exhibited cross reacting antibodies to other jellyfish, but no consistent pattern could be detected. 6. Significant titers were defined as those whose sera was positive after being be diluted 50-fold or greater. 7. Species specific IgM concentrations were regarded as significant only if their sera could be adsorbed against the homologous jellyfish antigen and the difference between adsorbed and non-adsorbed sera which were still positive was 50-fold. 8. Elevated persistent specific anti-jellyfish serum IgG concentrations which were still reactive if diluted 3000-fold were not protective against the cutaneous pain resulting from a natural sting.

Significant envenomation by Aurelia aurita, the moon jellyfish.

The case of a patient envenomated by Aurelia aurita, who developed significant local cutaneous lesions and immunospecific serum antibodies is reported. The lesions required more than ten days to heal. The patient developed significant cross-reacting antibodies to Chrysaora quinquecirrha antigens.

Jellyfish envenomation syndromes updated.

Jellyfish venoms are mixtures of toxic and/or antigenic polypeptides and enzymes pathogenic to human beings. As newer therapeutic agents become available to treat the various reactions to stings caused by these animals, an accurate diagnosis of the type of reaction the patient experiences and of the offending species will be necessary. Fatal reactions may be caused either by anaphylaxis or by the action of toxins in the venom on the heart, respiratory center, or kidneys. Cutaneous eruptions after envenomation may be local, generalized, exaggerated, recurrent, delayed, persistent, or occur at sites distant from the primary sting. Fat atrophy, pigmentary changes, vasospasm, and contractures with gangrene can occur after jellyfish stings. Identification of the envenoming animal can be made by actual visualization, examination for nematocysts on skin scraping, or serologically. It may also be predicted based on knowledge of location, time, and environmental circumstances of the encounter. First-aid measures designed to prevent additional nematocyst rupture are species-specific. Anaphylaxis should be prevented by the appropriate lifesaving measures. Other syndromes, caused by the toxins of the venom or mediated by humoral or cellular immune mechanisms, should be treated specifically.

Recurrent eruptions following unusual solitary coelenterate envenomations.

The case history of four patients is presented. The first patient exhibited normal immunologic reactions to large artificial intradermal challenge with jellyfish venom and later, multiple small natural stings. The second patient, presumably envenomated by a jellyfish, had four recurrent cutaneous eruptions in a linear configuration at the same anatomic site. Because her primary coelenterate contact occurred at a time when she was receiving systemic corticosteroids, it is assumed that the eruption due to the initial sting was delayed. The third and fourth patients exhibited recurrent eruptions after solitary envenomations by different coelenterates. These case histories demonstrate that multiple recurrent eruptions may follow solitary envenomations by different subphyla of coelenterates, that the initial eruption induced by the sting may be delayed by the administration of high doses of systemic corticosteroids, and that an immunologic reaction in both the B and T cell systems can follow jellyfish envenomation.

Effect of borohydride reduction on antibodies.

The effect of borohydride reducing reagents on monoclonal and polyclonal antibodies was examined by enzyme-linked immunosorbent assay (ELISA). Each antibody showed different stability characteristics to the reducing reagents. Sodium cyanoborohydride was at least five times milder toward immunological activity than sodium borohydride, however, sodium cyanoborohydride with a catalytic amount of metal ion (Zn2+ or Al3+) can be as harsh as sodium borohydride. Activated hydrophobic borohydrides, 9BBN-pyridine, did not have any advantages in respect to the stabilities of antibodies. Antibodies to be used for immunosorbent purification must be evaluated individually to determine whether their structure is stable to immobilization reagents and conditions prior to their linkage to the column support.

Venomous pelagic coelenterates: chemistry, toxicology, immunology and treatment of their stings.

Ten years have elapsed since our last review article on the toxicology of venomous pelagic coelenterates was published (Burnett and Calton, 1977). Investigation on important medusae and the chemistry of their nematocyst venoms have been expanding. The venomous jellyfish discussed here include the Portuguese man-o'war, (Physalia physalis), the sea nettle (Chrysaora quinquecirrha), the box jellyfish (Chironex fleckeri and/or Chiropsalmus quadrigatus), the cabbage head jellyfish (Stomolophus meleagris), the lion's mane jellyfish (Cyanea capillata), the Irukandji jellyfish (Carukia barnesi), the Moreton Bay Carybdeid medusa (Morbakka), and the mauve blubber (Pelagia noctiluca).