کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2066079 1076964 2007 7 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
CroFab™ total anti-venom activity measured by SE-HPLC, and anti-PLA2 activity assayed in vitro at physiological pH
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی بیوشیمی، ژنتیک و زیست شناسی مولکولی (عمومی)
پیش نمایش صفحه اول مقاله
CroFab™ total anti-venom activity measured by SE-HPLC, and anti-PLA2 activity assayed in vitro at physiological pH
چکیده انگلیسی

One problem in the development and refinement of anti-venoms is ascertaining both overall anti-venom reactivity and which key toxins are neutralized. Here we show by SE-HPLC that the in vitro reaction of CroFab™ anti-venin with Crotalus atrox venom asymptotically nears completion (>95%) by 11 min at 4 °C by following the change in area under chromatographic peaks. The peaks for reactants decrease and the formation of high molecular weight complexes increases with time. To assay the large number of samples a new microplate format phospholipase A2 (PLA2) assay at an initial pH of 7.5 was developed using phosphotidyl choline as the substrate. The change in absorbance is due to the pH change caused by release of fatty acids, and is linear with dilution of enzyme. This choice of substrate limits detection to PLA2 and nonspecific esterase (if any) activities. The neutralization mixtures show a dose dependent (CroFab™ anti-venin) inactivation of C. atrox PLA2 activity approaching a maximum of 85% neutralization. This approach of revealing antibody binding to venom components coupled with enzyme activity measurements is effective and may lead to greater in vitro assessment of antivenin activity in product development, and less routine use of mouse lethality assays.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Toxicon - Volume 49, Issue 6, May 2007, Pages 848–854
نویسندگان
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