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Mol. Biol. Cell 15, 1804-2818. Caspase-dependent regulation of histone deacetylase 4 nuclear-cytoplasmic shuttling promotes apoptosis. 2004

Paroni, G., Mizzau, M., Henderson, C., Del Sal, G., Schneider, C. and Brancolini, C.

Notes: In this study, the Anti-PARP p85 Fragment polyclonal antibody was used to confirm apoptosis in human IMR90 and MCF-7 cells by Western blotting lysates from UV- treated cells. The Anti-Cytochrome c monoclonal antibody was also used to immunocytochemically stain IMR90 cells that were transiently transfected with various mutant histone deacetylase-4 (HDAC4)-expressing constructs. Data from these experiments was expressed as the percent of cells showing cytochrome c staining in the cytosol. The researchers also mention using a TNT® Coupled Reticulocyte Lysate System to express and label various histone deacetylases (HDAC1, HDAC2, HDAC3, HDAC4, HDAC5, and HDAC6). The translation products were then used in in vitro cleavage assays with recombinant caspase-2 or caspase-3. (3174)

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Clin. Can. Res. 9, 2248-2259. Expression and Activation of the Nerve Growth Factor Receptor TrkA in Serous Ovarian Carcinoma 2003

Davidson, B., Reich, R., Lazarovici, P., Nesland, J.M., Skrede, M., Risberg, B., Trope, C.G., Florenes, V.A.

Notes: Anti-PARP p85 Fragment antibody was used to stain human peritoneal and pleural effusions. (2681)

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J. Biol. Chem. 278, 12579–12589. Role of caspases, Bid, and p53 in the apoptotic response triggered by histone deacetylase inhibitors trichostatin-A (TSA) and suberoylanilide hydroxamic acid (SAHA). 2003

Henderson, C., Mizzau, M., Paroni, G., Maestro, R., Schneider, C. and Brancolini, C.

Notes: The Anti-PARP p85 Fragment polyclonal antibody was used to detect cleaved p85 fragment PARP on Western blots of lysates from caspase-3 deficient human MCF-7 cells, and from cell lines MCF-7/C3WT or MCF-7/C3Cl with rescued caspase-3 expression. Anti-PARP p85 Fragment polyclonal antibody was also used on Western blots of lysates from stably transfected IMR90 human fibroblasts. A variety of treatments were used for inducing apoptosis including serum-free media, colchicine, daunorubicin, TSA (Trichostatin-A), and a compound called ET1 8OCH3. The researchers also used the Anti-Cytochrome c monoclonal antibody to immunocytochemically stain MCF-7/NEO cells. (3173)

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Clin. Can. Res. 8, 2035-2043. Validation of the pharmacodynamics of BMS-247550, an analogue of epothilone B, during a phase I clinical study 2002

McDaid, H.M., Mani, S., Shen, H-J., Muggia, F., Sonnichsen, D., Horwitz, S.B.

Notes: The authors investigated the mechanism of action of BMS-247550, a stable, synthetic derivative of epothilone B, in peripheral blood mononuclear cells and in breast tumor cells. Breast tumor cells expressing multidrug resistance were stained for tubulin bundle formation. Cytotoxicity in these biopsied cells also was examined by staining with Promega's Anti-PARP p85 Fragment, pAb. (2497)

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Cancer Res. 61, 318-326. Antiapoptotic role of endogenous nitric oxide in human melanoma cells. 2001

Salvucci, O., Carsana, M., Bersani, I., Tragni, G., and Anichini A.

Notes: The authors find that constitutive production of endogenous NO has an antiapoptotic function in human melanoma cells but not in normal melanocytes. To monitor apoptosis, Western blots were carried out with the Anti-PARP p85 Fragment pAb to monitor PARP cleavage. Melanoma cell extracts (10ug) were separated on 8% polyacrylamide minigels, and transferred to a PVDF membrane. Membranes were incubated with a 1:1000 dilution of the Anti-PARP p85 Fragment pAb, washed, and incubated with a biotinylated-anti rabbit IgG. Detection involved a streptavidin-horseradish peroxidase conjugate and a chemiluminescence substrate. Jurkat cells treated for 6 h with anti-FAS mAb were used as positive controls for Western blot detection of PARP cleavage (2354)

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Biotechniques 30, 886-891. Poly (ADP-ribose) polymerase cleavage monitored in situ in apoptotic cells. 2001

O'Brien, M.A., Moravec, R.A., and Riss T.L.

Notes: During apoptosis, Poly (ADP-ribose) polymerase (PARP) is cleaved by caspase-3 to generate an 85 kDa fragment (p85). An affinity-purified polyclonal antibody to the p85 fragment of PARP is characterized. In Western blot analysis with Jurkat cells treated with an anti-Fas antibody and with SH-Sy5Y cells treated with staurosporine, the antibody recognizes the 85-kDa (p85) fragment of PARP but not full-length PARP. The antibody was used at a concentration of 0.75 µg/ml for Western blots and at 2.5µg/ml for immunocytochemistry. TUNEL labeling of apoptotic Jurkat cells was performed with the DeadEnd™ Fluorometric TUNEL System. Immunocytochemistry was also performed using the Anti-β III Tubulin mAb. Good experimental details are given. (2355)

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J. Biol. Chem. 276(3), 1865-1872. Regulation of apoptosis by phosphatidylinositol 4,5-bisphosphate inhibition of caspases, and caspase inactivation of phosphatidylinositol phosphate 5-kinases 2001

Mejillano, M., Yamamoto, M., Rozelle, A.L., Sun, H.-Q., Wang, X., and Yin, H.L.

Notes: The Anti-PARP p85 Fragment pAb was used to assess apoptosis in human HEK293 cells transfected and overexpressing phosphatidylinositol phosphate 5-kinase and/or caspase-9. Less cleaved PARP was observed by Western blotting of extracts of cells expressing the kinase in addition to the caspase-9. (0019)

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Eur. J. Biochem. 267, 1083-1091. Changes in integrity and association of eukaryotic protein synthesis initiation factors during apoptosis. 2000

Bushell, M., Wood, W., Clemens, M.J., Morley, S.J.

Notes: The human BJAB Burkitt's lymphoma cell line was treated with 150ng/ml anti-Fas mAb (clone CH-11) for 4hr. Cell extracts were prepared and analyzed for cleavage products of translation initiation factors. To demonstrate that caspases were active and cleaving known substrates, the extracts were also analyzed for PARP cleavage with the Anti-PARP p85 Fragment pAb. PARP cleavage products were only obtained from cells treated with the anti-Fas mAb. No PARP cleavage was detected when the cells were treated with the caspase inhibitor Z-VAD-FMK concurrently with anti-Fas treatment. (1375)

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Exp. Cell Res. 255, 125-132. Cleavage of poly(ADP-ribose) polymerase measured in situ in individual cells: Relationship to DNA fragmentation and cell cycle position during apoptosis. 2000

Li, X. and Darzynkiewicz, Z.

Notes: The Anti-PARP p85 Fragment pAb was used in an extensive analysis of PARP cleavage relatively to DNA strand breaks induced in HL-60 cells. Apoptosis mediated by either campothecin or TNFα. PARP cleavage was detected by immunocytochemistry as well as multiparameter flow and laser scanning cytometry. PARP cleavage was readily apparent 30 minutes after TNFα treatment and PARP cleavage was apparent 90 minutes after campothecin treatment. The PARP cleavage was specific to S-phase of the cell cycle in the campothecin treated cells and was not specific to any cell phase during TNFα treatment. DNA strand breaks followed PARP cleavage by about 30 minutes. A strong correlation between DNA strand breaks and PARP cleavage is demonstrated. (0801)

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Mol. Cell 5, 981-992. NEMO/IKKgamma-deficient mice model incontinentia pigmenti. 2000

Schmidt-Supprian, M., Bloch, W., Courtois, G., Addicks, K., Israel, A., Rajewsky, and Pasparakis, M.

Notes: The Anti-PARP p85 Fragment pAb was used for immunohistochemical analysis of skin sections of wildtype and Nemo +/- mice. Analysis was performed on 5-10µm paraffin sections. (0018)

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Nat. Biotechnol. 18, 862-867. Reduction of liver Fas expression by an antisense oligonucleotide protects mice from fulminant hepatitis. 2000

Zhang, H., Cook, J., Nickel, J., Yu, R., Stecker, K., Myers, K., and Dean, N.M.

Notes: The authors downregulated the Fas ligand in mouse liver to determine the effect on Fas-mediated apoptosis. Apoptosis was monitored in mouse liver tissue by immunohistochemistry with the Anti-PARP p85 Fragment pAb. Cryostat sections (4.0 µm) were fixed in acetone and stained with a 1:100 of the anti-PARP antibody. An anti-rabbit HRP conjugate and DAB substrate was then used to visualize apoptotic cells. Slides were counterstained with hematoxylin. (2356)

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EMBO J. 19, 1157-1166. UV damage causes uncontrolled DNA breakage in cells from patients with combined features of XP-D and Cockayne syndrome. 2000

Berneburg, M., Lowe, J.E., Nardo, T., Araujo, S., Fousteri, M.I., Green, M.H.L., Krutmann, J., Wood, R.D., Stefanini, M. and Lehmann, A.R.

Notes: Primary human fibroblasts from normal and DNA-repair -deficient individuals were treated with UVB radiation for up to 72hr. Fibroblasts from Xeroderma pigmentosum patients all demonstrated cleavage of PARP at 72 hours. Normal fibroblasts had no detectable PARP cleavage. The results correlated well with TUNEL data. The cleavage of PARP was detected by Western blotting of cell extracts with the Anti-PARP p85 Fragment pAb. Excellent detail is provided. Human HL60 cells treated with 150mM campothecin for 4hr were used as positive controls. (1440)

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Science 287, 1500-1503. Virus-induced neuronal apoptosis blocked by the herpes simplex virus latency-associated transcript. 2000

Perng, G.-C., Jones, C., Ciacci-Zanella, J., Stone, M., Henderson, G., Yukht, A., Slanina, S.M., Hofman, F.M., Ghiasi, H., Nesburn, A.B., Wechsler, S.L.

Notes: Rabbits were infected with herpes viruses with or without the latency-associated transcript. The virus without the transcript produced more apoptotic neurons in the trigeminal ganglia than wildtype virus as judged by TUNEL staining and detection of the p85 fragment of PARP. Detection of the p85 Fragment was performed in situ with the Anti-PARP p85 Fragment pAb. Extensive work was performed to insure that Anti-PARP p85 Fragment staining correlated well with TUNEL staining by analyzing a series of sections. (0561)

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