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Biomedical subjects

H G Higashi

Publications and source records attributed to H G Higashi.

22 records · Page 2Linked to original sources
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Specific heterologous F(ab')2 antibodies revert blood incoagulability resulting from envenoming by Lonomia obliqua caterpillars.

Contact with Lonomia obliqua caterpillars results in a bleeding syndrome characterized by hemorrhage and blood coagulation disturbances. Conventional therapy using antifibrinolytics or cryoprecipitates has been unable to treat pathophysiologic alterations. As antivenoms are effective therapy for treatment of victims of venomous animals, a process of manufacturing a specific antilonomic serum by immunizing horses with Lonomia caterpillar bristle extracts (LBE) was developed. Lonomia caterpillar bristle extracts exhibited several protein bands on SDS-PAGE, induced blood coagulation abnormalities and lethality in mice, and stimulated specific antibody production in horses. Sera obtained from immunized horses were rich in anti-LBE specific antibodies distributed among the horse IgG isotypes. These antibodies had the ability to recognize various LBE antigens as well as to neutralize their coagulopathy-inducing activity. The antivenom manufactured by the developed process was composed of purified and sterilized F(ab')2 with ED50 = 38.61 microl, potency = 0.29 mg/ml, and 95% confidence limit of potency 0.20-1.36.

Animals↗
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Development and validation study for the chromatographic purification process for tetanus anatoxin on Sephacryl S-200 High Resolution.

The tetanus purified anatoxin is used in the preparation of multiple immunoprophylactics. WHO (World Health Organization) specifies that the tetanus anatoxin must exhibit a degree of purity greater than or equal to 1,000 Lf/mg protein nitrogen (PN). Today liquid chromatography is a well established technique for the purification of tetanus anatoxin and several different methods are used in production scale. On a small scale, we purified tetanus anatoxin on Sephacryl S-200 High Resolution (gel filtration) and we obtained a successful high-yield purification. On the basis of these results, by combining conventional tangential flow filtration (TFF) at 50,000 N.M.W.L. (Nominal Molecular Weight Limit) ultrafiltration membrane with gel filtration on Sephacryl S-200 High Resolution, we have been able to purify 14 lots of tetanus anatoxin using the Bioprocess System (Amersham Pharmacia Biotech) to a large scale operation. Using this method, 77,401,332 doses of tetanus toxoid were prepared in 14 consecutive lots, supporting the reproducibility and reliability of the method presented here.

Acrylic Resins↗
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Use of sephacryl high resolution (HR) in tetanus anatoxin purification.

The tetanus purified anatoxin is used in the preparation of the tetanus toxoid and multiple vaccines (dT, DT and DTP), all of them strictly following specifications established by the WHO with a minimum antigenic purity equal to 1,000 Lf/mgPN. Aiming to establish more sensitive and accurate methods for purification, samples from four different lots of tetanus anatoxin were submitted to gel filtration in twenty independent trials using the Sephacryl S-100 HR and S-200 HR resins. The Authors were careful to optimize their parameters of performance as to sample volume, elution and selectivity flow for tetanus anatoxin purification, allowing their use in industrial scale. The Sephacryl S-100 HR resin presented the best selectivity, that is, the best separation, allowing a greater linear-flow and, consequently, the best purity index. Satisfactory results were also achieved with the Sephacryl S-200 HR resin after optimization of chromatographic parameters for elution flow and volume of the sample applied. The good results of purification obtained, as well as the high chemical stability, have pointed out both the Sephacryl S-100 HR and S-200 HR resins as equally efficient for industrial production.

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