Western blot analysis of extracts from HeLa cells, untreated (-) or treated with Epothilone B (10 nM, 18 hr; +), using Phospho-Histone H3 (Ser10) (D2C8) XP® Rabbit mAb #3377 (upper) or Histone H3 (D1H2) XP® Rabbit mAb #4499 (lower).
Western blot analysis of extracts from HeLa cells, untreated (-) or treated with Epothilone B (10 nM, 18 hr; +), using Phospho-Aurora A (Thr288)/Aurora B (Thr232)/Aurora C (Thr198) (D13A11) XP® Rabbit mAb #2914 (upper) or Aurora A (1F8) Mouse mAb #12100 and COX IV (3E11) Rabbit mAb #4850 (lower).
Chemical structure of epothilone B.
Epothilone B is supplied as a lyophilized powder. For a 1 mM stock, reconstitute the 100 µg in 197 µl DMSO. Working concentrations and length of treatment can vary depending on the desired effect, but it is typically used at 10-1000 nM for 12-48 hr.
Store lyophilized or in solution at -20ºC, desiccated. Protect from light. In lyophilized form, the chemical is stable for 24 months. Once in solution, use within 3 months to prevent loss of potency. Aliquot to avoid multiple freeze/thaw cycles.
Epothilone A and B are taxol-like macrolides originally identified as antifungal, cytotoxic metabolites derived from the myxobacterium Sorangium cellulosum. Research studies demonstrate that epothilone B polymerizes tubulin into microtubules in vitro, which induces mitotic arrest at the G2/M phase and results in inhibition of cell proliferation and cytotoxicity (1-3). Cell cycle arrest at nanomolar IC50 values have been observed in many cell types, including HeLa (IC50 = 32 nM), Hs578T (IC50 = 3 nM) (3), as well as the multiple myeloma cell lines U266 and RPMI 8226 (IC50 = ~1-10 nM) (4). Investigations have shown that both epothilone A and B competitively inhibit binding of taxol to microtubules in vitro (3).
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