Monoclonal Antibody to Human Topoisomerase II Alpha: Western Blotting and Immunohistochemistry Reagent
A monoclonal antibody was prepared against an oligopeptide derived from the C-terminus of the p170 kDa human Topo IIa. The antibody was then affinity purified over a column containing the same peptide. The affinity purified antibody can be used to probe Western blots provided controls are included to clearly mark the position of the 170 kDa topo II. The reason for this is that human topo II is easily degraded or proteolyzed during routine extraction and manipulations; as a result, one may detect sub-bands. Heterogeneity in the SDS-PAGE pattern has been reported by several labs (Shelton et al., 1983; Halligan et al., 1985; Muller, et al., 1988) and recently proteolytic products of S. pombe topoisomerase II were reported (Shiozaki and Yanagida, 1991). Thus, the antibody may appear to be non-specific if there is extensive proteolysis. To counter this problem, a marker for human topoisomerase II is available (TG2011-3) which corresponds to the intact 170 kDa topoisomerase II. A second gene encoding han topoisomerase II (topo II-Beta or 180 kDa form) was reported (Chung et al., 1989; Tan et al., 1992). This antibody reacts specifically with the 170 kDa isoform of topo II.
Monoclonal Antibody Unit Definition:
The antibody concentration is in Western blotting units; one unit corresponds to a 1:1000 dilution of the antibody required to make a diluted probe for a routine Western blot. Thus, 10 units of antibody will make 10 ml of diluted probe (to be added directly to a single blot).
- Muller, M.T., Sptizner, J.R., DiDonato, J.D., Mehta, V.B., Tsutsui,K. and Tsutsui,K. 1988. Single stranded DNA cleavages by eukaryotic topoisomerase II. Biochemistry 27: 8369-8379.
- Halligan, B.D., Edwards, K.A. and Liu, L.F. 1985. Purification and characterization of a type II DNA topoisomerase from bovine calf thymus. J. Biolog. Chem. 260:2475-2482.
- Shelton,E.R., Osheroff, N. and Brutlag, D.L. 1983. DNA Topoisomerase II from Drosophila malanogaster. J. Biolog. Chem. 258:9530-9535.
- Chung,T., Drake,F., Tan,K., Per, S., Crooke, S., and Mirabelli,C. 1989. Characterization and immunological identification of cDNA clones encoding two human DNA topoisomerase II isozymes. Proc. Natl. Acad. Sci. U.S.A. 86:9431-9435.
- Shiozaki,K. and Yanagida,M. 1991. A functional 125 kDa core polypeptide of fission yeast DNA topoisomerase II. Mol. Cell. Biol. 11:6093-6102. °
Material Safety Data:
THIS PRODUCE IS NOT LICENSED OR APPROVED FOR ADMINISTRATION TO HUMANS OR TO ANIMALS OTHER THAN EXPERIMENTAL ANIMALS.
The product is sold as an antibody for research purposes only. Safe lab practices should be employed when handling this product.
Store the hydrated antibody at -20°C. Repeated freeze thaw cycles should be avoided, thus, we recommend storage in smaller aliquots. The diluted antibody can be re-used for probing blots, but should be kept at -20°C for long term storage or at 4°C for short periods.