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Description: ApopTag Plus Peroxidase In Situ Apoptosis Kit
Trade Name: ApopTag
Chemicon (Millipore)
Qty/Pk: 40 assays
Product Overview: The ApopTag® Plus Peroxidase In Situ Apoptosis Detection Kit detects apoptotic cells by labeling and detecting DNA strand breaks by the indirect TUNEL method. The kit provides sufficient reagents for immunoperoxidase staining of 40 samples, and includes ApopTag(R) Positive control Slides and DAB buffer and substrate. Results are visualized using brightfield microscopy.
The ApopTag® Peroxidase Kits have been qualified for use in histochemical and cytochemical staining of the following specimens: formalin-fixed, paraffin-embedded tissues, cryostat sections, cell suspensions, cytospins, and cell cultures. Whole mount-methods have been developed (34, 45). (See datasheet Sec. V. for References).
The staining specificity of the ApopTag® Peroxidase Kits has been demonstrated by Chemicon and many other laboratories. Chemicon has tested many types of model cell and tissue systems, including: (a) human prostate, thymus, and large intestine (in-house data); (b) rat ventral prostate post-castration (21), (c) rat thymus lymphocytes treated in vitro with dexamethasone (3, 13), (d) 14-day mouse embryo limbs (1) and (e) rat mammary gland in regression after weaning (36). In the thymocyte and prostate models, agarose gel electrophoresis was used to assess the amount of DNA laddering, which peaked coincidentally with the maximum percentage of stained cells. Numerous journal publications from laboratories worldwide have established the usefulness of ApopTag® Kits. (See datasheet Sec. V. References, Publications Citing ApopTag® Kits).View All » Background Information: Apoptosis is a form of cell death that eliminates compromised or superfluous cells. It is controlled by multiple signaling and effector pathways that mediate active responses to external growth, survival, or death factors. Cell cycle checkpoint controls are linked to apoptotic enzyme cascades, and the integrity of these and other links can be genetically compromised in many diseases, such as cancer. There are many books in print and hundreds of recent review articles about all aspects of apoptosis (e.g. 7, 11, 19, 24, 39, 42) and the methods for detecting it (e.g. 10, 32, 36).
Of all the aspects of apoptosis, the defining characteristic is a complete change in cellular morphology. As observed by electron microscopy, the cell undergoes shrinkage, chromatin margination, membrane blebbing, nuclear condensation and then segmentation, and division into apoptotic bodies which may be phagocytosed (11, 19, 24). The characteristic apoptotic bodies are short-lived and minute, and can resemble other cellular constituents when viewed by brightfield microscopy. DNA fragmentation in apoptotic cells is followed by cell death and removal from the tissue, usually within several hours (7). A rate of tissue regression as rapid as 25% per day can result from apparent apoptosis in only 2-3% of the cells at any one time (6). Thus, the quantitative measurement of an apoptotic index by morphology alone can be difficult.
DNA fragmentation is usually associated with ultrastructural changes in cellular morphology in apoptosis (26, 38). In a number of well-researched model systems, large fragments of 300 kb and 50 kb are first produced by endonucleolytic degradation of higher-order chromatin structural organization. These large DNA fragments are visible on pulsed-field electrophoresis gels (5, 43, 44). In most models, the activation of Ca2+- and Mg2+-dependent endonuclease activity further shortens the fragments by cleaving the DNA at linker sites between nucleosomes (3). The ultimate DNA fragments are multimers of about 180 bp nucleosomal units. These multimers appear as the familiar "DNA ladder" seen on standard agarose electrophoresis gels of DNA extracted from many kinds
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