These HR/NHEJ kits are cell based, meaning they have the HR or NHEJ GFP reporters pre-installed. A single plasmid (I-Sce1) is transfected to activate repair. Please see this table for a summary and comparison/application of each kit.
Transfection-based HR and NHEJ kits use transient transfections to make a pool of cells containing the DNA reporters to allow the investigator to interrogate the 2 major DNA repair pathways. The advantages are:
It uses GFP readout to measure the efficiency of repair.
There is zero background when repair is blocked or inhibited
The reporters for HR or NHEJ can be installed in virtually any cell line that can be transiently transfect
It is rapid and robust
It is ideal for drug screens to find novel inhibitors of HR/NHEJ
The kits are highly specific and can only report HR or NHEJ
Readout GFP can be done by FACS or Imaging
Entire pathway can be followed by live imaging in viable cells
Ideal to track repair kinetics at the single cell level
The marker set for topo II is used to unambiguously locate the position of the 170 kDa topo II in Western Blots. The human 170 kDa is present in SDS-PAGE loading buffer. Note that human topo II may break down spontaneously with prolonged storage or with repeated freezing and thawing cycles. TopoGEN, Inc. stipulates that the marker be 90% undegraded at the time of shipment.
This mouse monoclonal antibody is directed specifically against purified Topo IIb. Because the two isoforms are very close in MW, we demonstrate by band shifting analysis combined with Westerns that this antibody reacts predictably with the Beta isoform (Fig. 1).
This monoclonal antibody was raised to human topo I (TG2005H-RC1) using standard methods. The antibody (IgG class) works very well for Western blotting experiments. The epitope has not been mapped. This product has not been tested for IP or IF applications; however, given the high specificity seen on Westerns, it should work well in these applications.
Mouse monoclonal antibody prepared against an oligopeptide derived from the C-terminus of the p170 kDa human Topoisomerase II Alpha. The antibody was then affinity purified over a column containing the same peptide.
This monoclonal antibody was raised against purified recombinant protein and is specific for the C-terminal region (residues 748-976). The antibody is not ideal for use in western blotting experiments but will work on slot blot applications. It is well suited for immunohistochemistry with paraffin embedded tissue using 1:25 to 150 dilutions, the high temperature unmasking procedure and standard ABC techniques. It will work with frozen tissue although acetone fixation is recommended. We generally recommend 60 min incubation at 25 C for the primary antibody. Specific conditions used in each lab may vary as will specific applications for this AB. For…
The immunoglobulin fraction was purified from scleroderma patients and shown to react specifically with the 100 kDa form of Topo I using Western blotting. The antibody is particularly useful as a Western blotting probe and can be used at high dilutions (>1:5,000). Cross reactivity of this antibody with other species has been demonstrated (with calf thymus).
There are at least two type II topoisomerases in animal cells: Topo IIa and Topo IIb (Chung et al., 1989; Tan et al., 1992). The former is a cell cycle regulated enzyme of 170 kDa and the latter is not related to cell cycle phase and is slightly larger (180 kDa). This rabbit polyclonal antibody is directed specifically against the C-terminus of purified topo IIb.
This polyclonal antibody works with the following:
1
Immunohistochemistry Typical working dilution of 1:20 to 1:40 using a high temperature unmasking technique (see our technical literature). Typically, 60 min of primary AB incubation should be performed…
Rabbit Polyclonal Antibody to Human Topoisomerase II Alpha was prepared against a 16 residue oligopeptide derived from the carboxy terminal region of human topoisomerase II. The purity of the peptide was greater than 95% as determined by HPLC and mass spectral analysis. The antibody titer was determined by ELISA to be > 1:104. Immunoaffinity gel chromatography and antibody purification steps were carried out to obtain a homogeneously purified IgG.
A rabbit polyclonal antibody was prepared against a pool of oligopeptides derived from the carboxy terminal region of human topoisomerase IIIb. The purity of the peptides was greater than 95% as determined by HPLC and mass spectral analysis. Approximately 10 mg of purified peptide were coupled (through a terminal cysteine thiol) to KLH with a heterobifunctional cross-linking agent MBS at a ratio of 1 part peptide to 1 part KLH. The antibody titer was determined by ELISA to be > 1:10000. This antibody is ideal for Western blotting applications but has not been tested for other applications. The dilutions of…
This is a highly specific antibody, sold as crude serum (not affinity purified) that will neutralize enzymatic activity of yeast topoisomerase II. The antibody is specific for S. cerevisiae and does not cross react on Western blots with the human or calf thymus enzymes. Although this reagent may be suitable for use in immunofluorescent applications, it has not been characterized for such applications.
Antibody was raised to human topo I (purified from placenta) using New Zealand White rabbits. Serum was collected and processed using standard methods. The antibody can be used in Western blotting. Cross reactivity of this antibody with other species has not yet been tested. It is important to note that human topoisomerase I undergoes spontaneous degradation especially with repeat freezing/thawing. The activity of the enzyme is not affected significantly; however, sub-bands may be detectable in Western blotting experiments.
The Western blot marker set for topo I is used to unambiguously locate the position of the 100 kDa form of intact topo I. This product is supplied in SDS-PAGE loading buffer and ready to load directly onto an SDS-PAGE.
This vial contains a 16 residue oligopeptide derived from the carboxy terminal region of human topoisomerase IIa. The purity of the peptide was greater than 95% as determined by HPLC and mass spectral analysis. Approximately 10 mg of purified peptide were coupled (through a terminal cysteine thiol) to KLH with a heterobifunctional cross-linking agent MBS at a ratio of 1 part peptide to 1 part KLH. The peptide was used to immunize rabbits for antibody production. The resulting antibody reacts strongly and specifically with the 170 kDa Human Topo II protein. This peptide is used as a control reagent in…
TopoGEN has extensive experience with assays for topoisomerase inhibition in vivo. This analysis allows the investigator to ascertain whether a novel agent is active against endogenous Top2bin whole cells. An important benefit to this analysis is that one can use any tumor cell or tissue in order to establish clinical efficacy against a specific tumor cell line. We use essentially the same basic approach for Topo I, and Topo IIa,b. The kit is based upon ‘band shifting’ in a western blot using mono-specific antibody probes for Top2a (TG1024-A) or Top2b (TG1024-B.
This kit can be used to unambiguously detect a Top2a…
This assay kit contains all the reagents for cytolocalization of Human Top2a using immunofluorescence. The system is optimized for use with FITC or Rhodamine labeled secondary antibody with adherent cells. The method may work with other cells besides human. Mouse, rat and monkey cells will cross react with topo II antibody. The antibody is specific for p170 (topo II alpha).
When you become our customer, we become your partner in research. We review your data and offer valuable input. Feel free to contact us for further information about how we can assist your topoisomerase research endeavors.
