Technical Data
T8065-10X
Topoisomerase II alpha DNA (BSA & Azide Free)
Description:
The expression of topoisomerase II alpha is strongly restricted to proliferating cells. The topoisomerase II alpha antigen is preferentially expressed during G1, S, G2 and M phase of the cell cycle, while resting, non-cycling cells (G0 phase) lack topoisomerase II alpha. In addition, constantly proliferating cells (e.g. cell lines) react positively to topoisomerase II alpha during the entire cell-cycle. This specificity for proliferating cells might make it a useful tool for determination of the proliferative fraction in solid tumors such as mammary carcinomas and gangliomas. Note that Ki67/M1B1 is also present in G1-phase. Topo IIa is also implicated in drug resistance of tumor cells.

Topoisomerases change the topology of DNA via cleavage of single strand only. Suitable for use in DNA conformational analysis and topology as well as DNA repair, drug resistance, cell proliferation and leukemia studies. The double-helical configuration that DNA strands naturally reside in makes them difficult to separate, and yet they must be separated if enzymes are to transcribe the sequences that encode proteins, or if chromosomes are to be replicated. In so-called circular DNA, in which double helical segment is bent around and joined in a circle, the two strands are topologically linked, or knotted. They cannot be separated by any process that does not involve the breaking of strands. Topoisomerases catalyze and guide the unknotting of DNA.

Type I topoisomerases cut only one strand of DNA; type I topoisomerase of E. coli > E. coli (omega protein) relaxes negatively supercoiled DNA and does not act on positively supercoiled DNA. Type II topoisomerases cut both strands of DNA; type II topoisomerase of E. coli (DNA gyrase) increases the degree of negative supercoiling in DNA and requires ATP. It is inhibited by several antibiotics, including nalidixic acid and ovobiocin.

Applications and Suggested Dilutions:
ELISA (For coating, order Ab without BSA)
Immunoprecipitation: Use Protein A.10ug/mg protein lysate.
Western Blotting (Ab 5ug/ml for 2hrs at RT)
Immunofluorescence
Optimal dilution determined by the researcher.

Positive Control:
Raji cells or exponentially growing any cultured human cells.

Cellular Localization:
Nuclear

Molecular Weight of Antigen:
170kD

Storage and Stability:
May be stored at 4°C for short-term only. For long-term storage and to avoid repeated freezing and thawing, add glycerol (25–50%) and freeze at -20°C. Aliquots are stable for at least 12 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
TypeIsotypeCloneGrade
PabIgGPurified
SizeStorageShippingSourceHost
500ug4°C (-20°C Glycerol)Blue IceHumanRabbit
Concentration:
~1mg/ml
Immunogen:
KLH conjugate of a synthetic peptide, corresponding to aa 1513-1530 (Cys-RAKKPIKYLEESDEDDLF) from the C-terminus of human topoisomerase IIa.
Purity:
Purified by Protein A affinity chromatography.
Form
Supplied as a liquid in PBS, pH 7.4. Also supplied with BSA and Sodium Azide. See T8065-10.
Specificity:
Recognizes a 170kD protein, which is identified as topoisomerase IIa. It shows no crossreaction with Topoisomerase IIb or I. Specific for human.
Intended for research use only. Not for use in human, therapeutic, or diagnostic applications.
1. Wang JC. DNA topoisomerases. Ann Rev Biochem 54: 665-697, 1985.
2. Earnshaw WC, Halligan B, Cooke CA, Heck MMS, and Liu LF. Topoisomerse II is a structural component of mitotic chromosome scaffolds. J Cell Biol 100: 1706-1715, 1985.
3. Earnshaw WC and Heck MMS. Localization of topoisomerse II in mitotic chromosomes. J Cell Biol 100: 1716-1725, 1985.
4. Duguet M, Laverot C, harper H, Mirambeau G, and De Recondo AM. DNA topoisomerases from rat liver: physiological variations. Nucleic Acids Res 11: 1059-1075, 1983.
5. Tandou G, Mirambeau G, Lavenot C, der Garabedian A, Vermeersch J, and Duguet M. DNA topoisomerase activities in concanavalin A-stimualted lymphocytes. FEBS Lett 176: 431-435, 1984.
6. Heck MMS and Earnshaw WC. Topoisomerase II: a specific marker for cell proliferation. J Cell Biol 103: 2569-2581, 1986.
7. Heck MMS, Hittelman WN, and Earnshaw WC. Differential expression of DNA topoisomerases I and II during the eukaryotic cell cycle. Proc Natl Acad Sci (USA) 85: 1086-1090, 1988.
8. Verheijen R, Kuijpers HJH, Schlingemann RO, Boehmer ALM, Van Driel R, Brakenhoff GJ, and Ramaekers FCS. Ki-67 detects a nuclear matrix-associated proliferation-related antigen. I. Intracellular localization during interphase. J Cell Science 92: 123-130, 1989.
9. Verheijen R, Kuijpers HJH, Van Driel R, Beck JLM, Van Dierendonck JH, Brakenhoff GJ, and Ramaekers FCS. Ki-67 detects a nuclear matrix-associated proliferation-related antigen. II. Localization in mitotic cells and association with chromosomes. J Cell Science 92: 531-540, 1989.
10. Tandon AK, Hilsenbeck SG, Clark GM, Allred DC, Latham MD, Ross WE, and McGuire WL. Significnce of topoisomerase II in clinical breast cancer. Proc Am Assoc Cancer Res 32: 350 (Abstract 2076), 1991.
References Describing The Immuno-chemical Detection Of Topoisomerase II:

1. Caron PR; Watt P; Wang JC. The C-terminal domain of Saccharomyces cerevisiae DNA topoisomerase II. Molecular and Cellular Biology, 1994, 14(5):3197-207.
2. Chen G; Jaffrezou JP; Fleming WH; Duran GE; Sikic BI. Prevalence of multidrug resistance related to activation of the mdr1 gene in human sarcoma mutants derived by single-step doxorubicin selection. Cancer Research, 1994, 54(18):4980-7.
3. Daev E; Chaly N; Brown DL; Valentine B; Little JE; Chen X; Walker PR. Role of topoisomerase II in the structural and functional evolution of mitogen-stimulated lymphocyte nuclei. Experimental Cell Research, 1994, 214(1):331-42.
4. Ebert SN; Subramanian D; Shtrom SS; Chung IK; Parris DS; Muller MT. Association between the p170 form of human topoisomerase II and progeny viral DNA in cells infected with herpes simplex virus type 1. Journal of Virology, 1994, 68:1010-20.
5. Feldhoff PW; Mirski SE; Cole SP; Sullivan DM. Altered subcellular distribution of topoisomerase II alpha in a drug-resistant human small cell lung cancer cell line. Cancer Research, 1994, 54:756-62.
6. Khelifa T; Casabianca-Pignede MR; Rene B; Jacquemin-Sablon A. Expression of topoisomerases II alpha and beta in Chinese hamster lung cells resistant to topoisomerase II inhibitors. Molecular Pharmacology, 1994, 46(2):323-8.
7. Kubota N; Nishio K; Takeda Y; Ohmori T; Funayama Y; Ogasawara H; Ohira T; Kunikane H; Terashima Y; Saijo N. Characterization of an etoposide-resistant human ovarian cancer cell line. Cancer Chemotherapy and Pharmacology, 1994, 34(3):183-90.
8. Neri LM; Santi S; Marugg RA; Riederer BM; Capitani S; Cataldi A; Martelli AM. In vitro heat exposure induces a redistribution of nuclear matrix proteins in human K562 erythroleukemia cells. Experimental Cell Research, 1994, 213(1):275-85.
9. Yang JM; Goldenberg S; Gottesman MM; Hait WN. Characteristics of P388/VMDRC.04, a simple, sensitive model for studying P-glycoprotein antagonists. Cancer Research, 1994, 54(3):730-7.
10. Zini N; Santi S; Ognibene A; Bavelloni A; Neri LM; Valmori A; Mariani E; Negri C; Astaldi-Ricotti GC; Maraldi NM. Discrete localization of different DNA topoisomerases in HeLa and K562 cell nuclei and subnuclear fractions. Experimental Cell Research, 1994, 210(2):336-48.
11. Ishikawa H; Kawano MM; Okada K; Tanaka H; Tanabe O; Sakai A; Asaoku H; Iwato K; Nobuyoshi M; Kuramoto A. Expressions of DNA topoisomerase I and II gene and the genes possibly related to drug resistance in human myeloma cells. British Journal of Haematology, 1993, 83(1):68-74.
12. Kuwana M; Kaburaki J; Okano Y; Inoko H; Tsuji K. The HLA-DR and DQ genes control the autoimmune response to DNA topoisomerase I in systemic sclerosis (scleroderma). Journal of Clinical Investigation, 1993, 92(3):1296-301.
13. Mann K. Topoisomerase activity is associated with purified SV40 T antigen. Nucleic Acids Research, 1993, 21(8):1697-704.
14. McPherson SM; Longo FJ. Nicking of rat spermatid and spermatozoa DNA: possible involvement of DNA topoisomerase II. Developmental Biology, 1993, 158(1):122-30.
15. Negri C; Scovassi AI; Braghetti A; Guano F; Astaldi Ricotti GC. DNA topoisomerase II beta: stability and distribution in different animal cells in comparison to DNA topoisomerase I and II alpha. Experimental Cell Research, 1993, 206(1):128-33.
16. Prosperi E; Stivala LA; Sala E; Scovassi AI; Bianchi L. Proliferating cell nuclear antigen complex formation induced by ultraviolet irradiation in human quiescent fibroblasts as detected by immunostaining and flow cytometry. Experimental Cell Research, 1993, 205(2):320-5.
17. Robinson RG; Rapp L; Bowdish KJ; Graham MA; Huff AC; Coughlin SA. Isolation and characterization of monoclonal antibodies to a recombinant human topoisomerase II polypeptide. Hybridoma, 1993, 12(4):407-15.
18. Scovassi AI; Mariani C; Negroni M; Negri C; Bertazzoni U. ADP-ribosylation of nonhistone proteins in HeLa cells: modification of DNA topoisomerase II. Experimental Cell Research, 1993, 206(1):177-81.
19. Taagepera S; Rao PN; Drake FH; Gorbsky GJ. DNA topoisomerase II alpha is the major chromosome protein recognized by the mitotic phosphoprotein antibody MPM-2. Proceedings of the National Academy of Sciences of the United States of America, 1993, 90(18):8407-11.
20. Tuccari G; Rizzo A; Giuffre G; Barresi G. Immunocytochemical detection of DNA topoisomerase type II in primary breast carcinomas: correlation with clinico-pathological features. Virchows Archiv. a, Pathological Anatomy and Histopathology, 1993, 423(1):51-5.
21. Volm M; Kastel M; Mattern J; Efferth T. Expression of resistance factors (P-glycoprotein, glutathione S-transferase-pi, and topoisomerase II) and their interrelationship to proto-oncogene products in renal cell carcinomas. Cancer, 1993, 71(12):3981-7.
22. Efferth T; Mattern J; Volm M. Immunohistochemical detection of P glycoprotein, glutathione S transferase and DNA topoisomerase II in human tumors. Oncology, 1992, 49(5):368-75.
23. Ferguson PJ; Garland A; Lampe HB. Measurement of proliferative index in squamous cell carcinoma by flow cytometry. Journal of Otolaryngology, 1992, 21(5):331-8.
24. Friche E; Danks MK; Beck WT. Characterization of tumor cell resistance to 4'-deoxy-4'-iododoxorubicin developed in Ehrlich ascites cells in vivo. Cancer Research, 1992, 52(20):5701-6.
25. Gekeler V; Frese G; Noller A; Handgretinger R; Wilisch A; Schmidt H; Muller CP; Dopfer R; Klingebiel T; Diddens H; et al. Mdr1/P-glycoprotein, topoisomerase, and glutathione-S-transferase pi gene expression in primary and relapsed state adult and childhood leukaemias. British Journal of Cancer, 1992, 66(3):507-17.
26. Goswami PC; Hill M; Higashikubo R; Wright WD; Roti Roti JL. The suppression of the synthesis of a nuclear protein in cells blocked in G2 phase: identification of NP-170 as topoisomerase II. Radiation Research, 1992, 132(2):162-7.
27. Holden JA; Rolfson DH; Wittwer CT. The distribution of immunoreactive topoisomerase II Protein in human tissues and neoplasms. Oncology Research, 1992, 4:157-66.
28. Negri C; Chiesa R; Cerino A; Bestagno M; Sala C; Zini N; Maraldi NM; Astaldi Ricotti GC. Monoclonal antibodies to human DNA topoisomerase I and the two isoforms of DNA topoisomerase II: 170-and 180-kDa isozymes. Experimental Cell Research, 1992, 200(2):452-9.
29. Prosperi E; Sala E; Negri C; Oliani C; Supino R; Astraldi Ricotti GB; Bottiroli G. Topoisomerase II alpha and beta in human tumor cells grown in vitro and in vivo. Anticancer Research, 1992, 12:2093-9.
30. Takigawa N; Ohnoshi T; Ueoka H; Kiura K; Kimura I. Establishment and characterization of an etoposide-resistant human small cell lung cancer cell line. Acta Medica Okayama, 1992, 46(3):203-12.
31. Volm M; Mattern J. Expression of topoisomerase II, catalase, metallothionein and thymidylate-synthase in human squamous cell lung carcinomas and their correlation with doxorubicin resistance and with patients’ smoking habits. Carcinogenesis, 1992, 13(11):1947-50.
32. Volm M; Mattern J; Efferth T; Pommerenke EW. Expression of several resistance mechanisms in untreated human kidney and lung carcinomas. Anticancer Research, 1992, 12(4):1063-7.
33. Zini N; Martelli AM; Sabatelli P; Santi S; Negri C; Astaldi Ricotti GC; Maraldi NM. The 180-kDa isoform of topoisomerase II is localized in the nucleolus and belongs to the structural elements of the nucleolar remnant. Experimental Cell Research, 1992, 200(2):460-6.
34. Cole SP; Chanda ER; Dicke FP; Gerlach JH; Mirski SE. Non-P-glycoprotein-mediated multidrug resistance in a small cell lung cancer cell line: evidence for decreased susceptibility to drug-induced DNA damage and reduced levels of topoisomerase II. Cancer Research, 1991, 51(13):3345-52.
35. Kaufmann SH; McLaughlin SJ; Kastan MB; Liu LF; Karp JE; Burke PJ. Topoisomerase II levels during granulocytic maturation in vitro and in vivo. Cancer Research, 1991, 51(13):3534-43.
36. Sugawara I; Iwahashi T; Okamoto K; Sugimoto Y; Ekimoto H; Tsuruo T; Ikeuchi T; Mori S. Characterization of an etoposide-resistant human K562 cell line, K/eto. Japanese Journal of Cancer Research, 1991, 82(9):1035-43.
37. Chen YN; Mickley LA; Schwartz AM; Acton EM; Hwang JL; Fojo AT. Characterization of adriamycin-resistant human breast cancer cells which display overexpression of a novel resistance-related membrane protein. Journal of Biological Chemistry, 1990, 265(17):10073-80.
38. Kato S; Ideguchi H; Muta K; Nishimura J; Nawata H. Mechanisms involved in the development of adriamycin resistance in human leukemic cells. Leukemia Research, 1990, 14(6):567-73.
39. Smith PJ; Makinson TA. Cellular consequences of overproduction of DNA topoisomerase II in an ataxia-telangiectasia cell line. Cancer Research, 1989, 49(5):1118-24.
40. Ferguson PJ; Fisher MH; Stephenson J; Li DH; Zhou BS; Cheng YC. Combined modalities of resistance in etoposide-resistant human KB cell lines. Cancer Research, 1988, 48(21):5956-64.
41. Heck MM; Hittelman WN; Earnshaw WC. Differential expression of DNA topoisomerases I and II during the eukaryotic cell cycle. Proceedings of the National Academy of Sciences of the United States of America, 1988, 85(4):1086-90.
42. Hsiang YH; Wu HY; Liu LF. Proliferation-dependent regulation of DNA topoisomerase II in cultured human cells. Cancer Research, 1988, 48(11):3230-5.
43. Bapat AR; Han FS; Liu ZC; Zhou BS; Cheng YC. Studies on DNA topoisomerases I and II in herpes simplex virus type 2-infected cells. Journal of General Virology, 1987, 68:2231-7.
44. Per SR; Mattern MR; Mirabelli CK; Drake FH; Johnson RK; Crooke ST. Characterization of a subline of P388 leukemia resistant to amsacrine: evidence of altered topoisomerase II function. Molecular Pharmacology, 1987, 32(1):17-25.
45. Gasser SM; Laroche T; Falquet J; Boy de la Tour E; Laemmli UK. Metaphase chromosome structure. Involvement of topoisomerase II. Journal of Molecular Biology, 1986, 188:613-29.
46. Heck MM; Earnshaw WC. Topoisomerase II: A specific marker for cell proliferation. Journal of Cell Biology, 1986, 103:2569-81.
47. Earnshaw WC; Halligan B; Cooke CA; Heck MM; Liu LF. Topoisomerase II is a structural component of mitotic chromosome scaffolds. Journal of Cell Biology, 1985, 100(5):1706-15.
48. Earnshaw WC; Heck MM. Localization of topoisomerase II in mitotic chromosomes. Journal of Cell Biology, 1985, 100:1716-25.