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Biomedical subjects

H G Higashi

Publications and source records attributed to H G Higashi.

At least 19 recordsLinked to original sources
PubMed2007

Determination of low tetanus or diphtheria antitoxin titers in sera by a toxin neutralization assay and a modified toxin-binding inhibition test.

A method for the screening of tetanus and diphtheria antibodies in serum using anatoxin (inactivated toxin) instead of toxin was developed as an alternative to the in vivo toxin neutralization assay based on the toxin-binding inhibition test (TOBI test). In this study, the serum titers (values between 1.0 and 19.5 IU) measured by a modified TOBI test (Modi-TOBI test) and toxin neutralization assays were correlated (P < 0.0001). Titers of tetanus or diphtheria antibodies were evaluated in serum samples from guinea pigs immunized with tetanus toxoid, diphtheria-tetanus or triple vaccine. For the Modi-TOBI test, after blocking the microtiter plates, standard tetanus or diphtheria antitoxin and different concentrations of guinea pig sera were incubated with the respective anatoxin. Twelve hours later, these samples were transferred to a plate previously coated with tetanus or diphtheria antitoxin to bind the remaining anatoxin. The anatoxin was then detected using a peroxidase-labeled tetanus or diphtheria antitoxin. Serum titers were calculated using a linear regression plot of the results for the corresponding standard antitoxin. For the toxin neutralization assay, L+/10/50 doses of either toxin combined with different concentrations of serum samples were inoculated into mice for anti-tetanus detection, or in guinea pigs for anti-diphtheria detection. Both assays were suitable for determining wide ranges of antitoxin levels. The linear regression plots showed high correlation coefficients for tetanus (r(2) = 0.95, P < 0.0001) and for diphtheria (r(2) = 0.93, P < 0.0001) between the in vitro and the in vivo assays. The standardized method is appropriate for evaluating titers of neutralizing antibodies, thus permitting the in vitro control of serum antitoxin levels.

Animals↗
PubMed2005

The improvement of the therapeutic anti-Lachesis muta serum production in horses.

The main features associated with pit viper envenomations include the intense local lesions such as oedema, necrosis, acute renal failure and other effects. The severity of these reactions to snakebite depends on the degree of envenomation. Lachesis muta venom (LMV) has weak lethal activity, but due to the large amount often inoculated, the effects are extremely severe and demand anti-venom with a high neutralizing capacity. LMV had the lowest neutralizing antibody induction capacity in horses when compared with that of other venoms. For example, Bothrops anti-venom serum neutralizes 180 times the equivalent LD(50) to Bothrops venom; Crotalus anti-venom neutralizes 250 LD(50) of this venom, while Lachesis anti-venom neutralizes only five LD(50) of the Lachesis toxins. To examine the reasons for this low antibody induction, the H(GP) mouse line, genetically selected for high antibody production received, at different times during immunization with sheep erythrocytes (SE), whole LMV and isolated venom fractions I-VI eluted by gel-filtration chromatography on Superdex75. The specific antibody responsiveness showed a partial, but significant suppression of the anti-SE antibody responses during the kinetics of the primary and even the secondary immunizations, after 50-100 microg of fractions IV and V administration 72-48 h before the first antigen injections. Fraction IV was then applied in a Superose 12 column and three samples were obtained. The peak IVA containing a component of Mr 27 kDa was liable with the immunosuppressive effect as made evident by its effect on the H mice anti-SE responses. Horses receiving the LMV exempt of fractions IV and V produce highly significant anti-Lachesis sera with a 45 LD(50) neutralizing activity, providing, for the first time, an efficient specific therapeutic heterologous serum for human use.

Animals↗
PubMed2001

Cross reactivity of different specific Micrurus antivenom sera with homologous and heterologous snake venoms.

Coral snakes are the only Elapids in America. They are represented by three genera: Leptomicrurus, Micruroides and Micrurus, of which the latter are the most abundant and diversified group. Little is known about the biochemistry of Micrurus venoms due to low availability. Here, we present a study on the cross reactivity of different specific Micrurus antivenom with homologous and heterologous snake venoms in order to contribute to the generation of more efficient antiserum for therapeutic purposes. The three specific antisera tested, anti-Micrurus corallinus, anti-Micrurus frontalis, and anti-Micrurus spixii, as well as the bivalent anti-elapid venom sera, raised against a mixture (50% each) of Micrurus frontalis and Micrurus corallinus venoms, were assayed by Western Blot against Micrurus and non-Micrurus elapid venoms. An antisera raised against a recombinant alpha-neurotoxin-like protein from Micrurus corallinus venom, only reacted in Western blot with its homologous venom, indicating that this protein is specific for Micrurus corallinus coral snake.

Animals↗
PubMed2000

Neutralization of crotaline snake venoms from Central and South America by antivenoms produced in Brazil and Costa Rica.

A study was performed on the ability of antivenoms, produced in Brazil and Costa Rica, to neutralize lethal, hemorrhagic and coagulant activities of the venoms of 16 species of Central and South American snakes of the subfamily Crotalinae. Neutralization of lethality was studied by two different methods routinely used in the quality control of antivenoms at Instituto Butantan (IB) and Instituto Clodomiro Picado (ICP). Both antivenoms neutralized the majority of the venoms studied, but the values of effective doses 50% (ED(50)) differed markedly depending on the method used. In general, higher potencies were obtained with the method of ICP, where a challenge dose corresponding to 4 LD(50)s is used, than with the method of IB, where a challenge dose of 5 LD(50)s is employed. All venoms induced hemorrhagic activity in the mouse skin test, which was effectively neutralized by the two antivenoms. All venoms, except those of Porthidium nasutum and Bothriechis lateralis, induced coagulation of human plasma in vitro and both antivenoms were effective in the neutralization of this activity. In conclusion, our results provide evidence of an extensive cross reactivity between these antivenoms and Central and South American crotaline snake venoms.

Animals↗
PubMed1998

A randomized, blinded, comparative trial of one pepsin-digested and two whole IgG antivenoms for Bothrops snake bites in Uraba, Colombia. The Regional Group on Antivenom Therapy Research (REGATHER).

The therapeutic efficacy and the incidence of early antivenom reactions (EARs) were compared in a clinical trial performed in 79 patients bitten by Bothrops sp. in Urabá, Colombia. Patients were randomized into three groups according to the antivenom administered: A (n = 30, Butantan polyspecific, pepsin-digested Bothrops antivenom); B (n = 27, Butantan polyspecific, whole IgG Bothrops antivenom); and C (n = 22, Colombian commercial, monovalent, whole IgG Bothrops antivenom). The groups were comparable in all clinical and epidemiologic aspects; 33 patients had mild, 22 moderate, and 24 severe envenoming. At the doses used (two, four, and six vials [10 ml/vial] for mild, moderate, and severe envenomings, respectively) there were no differences between the antivenoms in restoring normal hemostatic parameters within 24 hr. The evolution of local envenoming was comparable in the three groups. Serum venom/antivenom kinetics determined by ELISA showed a complete clearance of venom levels 1 hr after treatment in mild/moderate envenomings. In severe cases, venom levels remained detectable up to 24 hr and recurrence of antigenemia was observed in some cases. Antivenom concentrations remained at high levels up to 24 hr of treatment. The incidence of EARs was significantly different in the groups: A (36.7%), B (11.1.%), and C (81.8%). There were no life-threatening anaphylactic reactions. We conclude that the efficacy of the three antivenoms was similar in neutralizing human Bothrops envenomings and that the production of whole IgG antivenoms by caprylic acid fractionation is a good alternative for reducing the incidence of EARs.

Adolescent↗
PubMed1996

Development of an antivenom against toxins of Lonomia obliqua caterpillars.

Skin contact with caterpillars of Lonomia moths causes haemostatic disorders that may evolve into a haemorrhagic syndrome. Replacement therapy has been shown to exacerbate the clinical symptoms of this envenoming. In this study it is shown that horses immunized with a bristle extract of L. obliqua caterpillars produced IgG antibodies that completely neutralized, in vitro, the toxin(s) responsible for the blood incoagulability observed in rats. This antivenom offers the possibility of specific treatment for envenoming caused by contact with caterpillars of Lonomia moths.

Animals↗
PubMed1995

Antigenic cross-reactivity among components of Brazilian Elapidae snake venoms.

Snake venoms from M. corallinus (LD50 = 7.1 +/- 0.83 micrograms), M. frontalis (LD50 = 19.3 +/- 3.13 micrograms), M. ibiboboca (LD50 = 19.8 +/- 2.07 micrograms) and M. spiixi (LD50 = 6.7 +/- 1.25 micrograms) (family Elapidae, genus Micrurus) injected into horses alone or in combination (M. corallinus with M. frontalis) elicit antibody production, as indicated in vivo by neutralization of venom lethality and in vitro by enzyme-linked immunosorbent assay (ELISA), immunoelectrophoresis (IE) and Western blotting (WB). Venom lethality was efficiently neutralized by the antisera, with the monovalent antivenoms being more efficient than the bivalent antivenom. Antibodies against venom components were detected by all antisera at different titers by ELISA. Upon IE, antisera against M. spiixi and M. frontalis venoms cross-reacted with the four types of venoms studied and recognized several molecular components, the precipitin lines obtained had distinct intensities and electrophoretic motilities, whereas the antivenom against M. corallinus only recognized components of its venom but not of the others. All antivenoms cross-reacted with all the elapid venoms in WB revealing several bands with distinct MWs in M. corallinus and M. spiixi venoms, two very sharp and separate bands in M. corallinus venom and a very sharp band of high MW together with several other smaller and faint bands in M. frontalis venom. The data indicate that snake venoms of the genus Micrurus are good immunogens that contain many cross-reactive molecules, and that their toxic components are neutralized more effectively by monovalent rather than by bivalent antivenom.

Animals↗
PubMed1994

An effective toxoid from tetanus toxin prepared by stepwise iodination of purified toxin.

Purified samples of tetanus toxin were gradually iodinated by stepwise addition of iodine, up to saturation. The residual capacity of each sample to provoke tetanus was tested by injecting sc 25 Lf (Limit of flocculation) in mice. Toxicity diminished in relation to iodine incorporated, and with iodine-saturated samples, doses of up to 100 Lf in mice, or 500 Lf in guinea-pigs, proved innocuous. Mice immunized with two doses of this toxoid adsorbed on Al(OH)3, and challenged with standard lethal toxin, gained protection against 10 MLD (Minimum Lethal Doses). Guinea-pigs were immunized by a single sc dose of 27.5 Lf with fluid toxoid, and all resisted challenge against 10 MLD applied 30 days later. The mice sera gave strong immunoprecipitation lines against the native toxin. The findings indicated that by controlled iodination of tetanus toxin an effective and inexpensive toxoid can be prepared.

Animals↗
PubMed1994

Antigenic cross-reactivity among the venoms from several species of Brazilian scorpions.

The venoms of seven species of scorpions living in different regions of Brazil were analysed with regard to their lethality, antigenic cross-reactivity and ability to induce antibody production. In mice, the tested scorpion venoms can be grouped as: (a) highly toxic: Tityus stigmurus Thorell (LD50 = 0.773 mg/kg), Tityus bahiensis (Perty) (LD50 = 1.062 mg/kg), Tityus serrulatus Lutz and Mello (LD50 = 1.160 mg/kg), and Tityus costatus (Karsch) (LD50 = 1.590 mg/kg); (b) moderately toxic: Tityus cambridgei Pocock (LD50 = 12.136 mg/kg); and (c) practically nontoxic: Rhopalurus agamemnon (Koch) (LD50 = 36.363 mg/kg), and Brotheas amazonicus Lourenço (LD50 = 90.909 mg/kg). On electrophoresis the venoms showed many protein bands displayed along the chromatogram, most of them cross-reacting in immunoelectrophoresis and immunoblotting using horse anti-T. serrulatus, anti-T. bahiensis or anti-T. serrulatus+T. bahiensis sera as probes. The antibodies present in these antivenoms combine with venom components as measured in vitro by the ELISA assay, and neutralize their lethal effects in vivo. These results indicate that horse anti-venoms against a mixture of T. serrulatus and T. bahiensis venoms or only against T. serrulatus venom yield an antibody population able to neutralize the toxic effects found in all venoms studied.

Animals↗
PubMed1992

Use of enzyme immunoassays to compare the effect and assess the dosage regimens of three Brazilian Bothrops antivenoms. The Butantan Institute Antivenom Study Group (BIASG).

The effect of the three main Brazilian polyspecific antivenoms on venom clearance was assessed in 118 moderately envenomed victims of bites by Bothrops species (mainly B. jararaca) in Sao Paulo State, Brazil. Serum samples taken from patients at intervals during their stay in the hospital and at followup approximately four weeks later were tested by enzyme immunoassay for the presence of whole venom and therapeutic antivenom. Results indicated that in patients treated with the standard regimen of either four (40 ml) or eight (80 ml) ampules of each antivenom, venom was cleared from the circulation within four days of antivenom administration. However, high concentrations of antivenom persisted for approximately 10 days and remained detectable until 30-50 days after administration. This suggests that patients may be being treated with excessive amounts of antivenom in Brazil. This practice increases the national cost of antivenom therapy and may contribute to the high frequency of antivenom reactions. Clinically, there was no obvious difference in the efficacy between the three antivenoms.

Adolescent↗
PubMed1989

Immunization of equines with phospholipase A2 protects against the lethal effects of Crotalus durissus terrificus venom.

Equines (2 horses and 2 donkeys) immunized with whole Crotalus durissus terrificus venom or its phospholipase A2 component either presented an increased survival time determined 3 days after challenge or were totally resistant to a challenging lethal dose of 200 mg crude venom 270 days after the initial immunization or 90 days after the last booster injection. The resistance was demonstrable on the basis of a good correlation with antibody titers determined by the ELISA method but not with the flocculation and neutralization assays. Since phospholipase A2 is essentially nontoxic, it can be used as a substitute for whole venom for the production of commercial antisera and as an immunizing agent in prophylactic programs.

Animals↗
PubMedDate not supplied

Effect of beta-propiolactone treatment on the complement activation mediated by equine antisera.

Reduction of complement activation through an alteration of the Fc fragment of immunoglobulins by beta-propiolactone treatment was carried out in equine antisera raised against rabies virus, Bothrops venoms and diphtherial toxin. Results were evaluated by means of an anaphylactic test performed on guinea-pigs, and compared to the ones obtained with the same sera purified by saline precipitation (ammonium sulfate), followed or not by enzymatic digestion with pepsin. Protein purity levels for antibothropic serum were 184.5 mg/g and 488.5 mg/g in beta-propiolactone treated and pepsin-digested sera, respectively. The recovery of specific activity was 100% and 62.5% when using antibothropic serum treated by beta-propiolactone and pepsin digestion, respectively. The antidiphtherial and anti-rabies sera treated with beta-propiolactone and pepsin presented protein purity levels of 5,698 and 7,179 Lf/g, 16,233 and 6,784 IU/g, respectively. The recovery of specific activity for these antisera were 88.8%, 77.7%, 100% and 36.5%, respectively. beta-propiolactone treatment induced a reduction in complement activation, tested "in vivo", without significant loss of biological activity. This treatment can be used in the preparation of heterologous immunoglobulins for human use.

Animals↗