Tissue specimens from mice were formalin-fixed then subjected to IHC staining as described previously [27 (link),34 (link)]. Briefly, the tissue sections were de-waxed and rehydrated. The sections were stained with monoclonal antibodies against cleaved PARP and cleaved caspase3 (Cell Signaling) for 24 h at 4°C. After washing, the samples were probed with peroxidase-labeled goat anti-rabbit secondary antibody (Epitomics, Burlinggame, CA) and detected with an ABC kit (Vector Laboratories, Burlingame, CA).
Cleaved parp and cleaved caspase3
Manufactured by Cell Signaling Technology
Sourced in United States
Cleaved PARP and Cleaved Caspase-3 are laboratory reagents used to detect apoptosis, a programmed cell death process. Cleaved PARP is a fragment of the Poly(ADP-ribose) Polymerase enzyme, while Cleaved Caspase-3 is a fragment of the Caspase-3 enzyme. Both of these fragments are generated during apoptosis and serve as markers for this cellular event.
Automatically generated - may contain errors
Lab products found in correlation
Peroxidase labeled goat anti rabbit secondary antibody, by Abcam (1 mentions)
Abc kit, by Vector Laboratories (1 mentions)
Apoptotic dna ladder kit, by Roche (1 mentions)
Annexin 5 fitc, by BD (1 mentions)
Trypan blue solution, by Thermo Fisher Scientific (1 mentions)
Cell culture media, by Thermo Fisher Scientific (1 mentions)
Propidium iodide, by BD (1 mentions)
Clarity western ecl substrate, by Bio-Rad (1 mentions)
Serum supreme, by Lonza (1 mentions)
Amersham ecl western blotting, by Cytiva (1 mentions)
3 protocols using cleaved parp and cleaved caspase3
1
Immunohistochemical Analysis of Apoptosis Markers
2
Synthesis and Characterization of Ruthenium Complexes
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Ru(bathophenanthroline)3 (1 ) and Ru(bathophenanthroline disulfonate)3 (2 ) were synthesized using previously established
procedures.20 (link),44 All cell lines were purchased
from ATCC. Cell culture media, heat-inactivated
fetal bovine serum (FBS), 4–20% tris-glycine precast gels,
Dulbecco’s phosphate buffered saline (DPBS), penicillin-streptomycin
solution (pen-strep), and 0.4% Trypan Blue solution were from Invitrogen.
35 mm wide, 4-compartment CELLview cell culture dishes were obtained
from USA Scientific. Serum supreme was from Lonza. Hoechst 33342,
Lysotracker Green DND-26 and Mitotracker Green FM were purchased from
Invitrogen. Propidium iodide (PI) and FITC-Annexin V were obtained
from BD Science. Trimethylrhodamine ethyl ester (TMRE) was purchased
from Sigma–Aldrich. Antibodies for PARP-1, procaspase 3, and
GAPDH were from Santa Cruz Biotechnology, Inc., while cleaved PARP
and cleaved caspase 3 was from Cell Signaling Technology. RIPA buffer
was purchased from Santa Cruz Biotechnology, Inc. Clarity Western
ECL Substrate was from Bio-Rad. An apoptotic DNA-ladder kit was purchased
from Roche Applied Science.
procedures.20 (link),44 All cell lines were purchased
from ATCC. Cell culture media, heat-inactivated
fetal bovine serum (FBS), 4–20% tris-glycine precast gels,
Dulbecco’s phosphate buffered saline (DPBS), penicillin-streptomycin
solution (pen-strep), and 0.4% Trypan Blue solution were from Invitrogen.
35 mm wide, 4-compartment CELLview cell culture dishes were obtained
from USA Scientific. Serum supreme was from Lonza. Hoechst 33342,
Lysotracker Green DND-26 and Mitotracker Green FM were purchased from
Invitrogen. Propidium iodide (PI) and FITC-Annexin V were obtained
from BD Science. Trimethylrhodamine ethyl ester (TMRE) was purchased
from Sigma–Aldrich. Antibodies for PARP-1, procaspase 3, and
GAPDH were from Santa Cruz Biotechnology, Inc., while cleaved PARP
and cleaved caspase 3 was from Cell Signaling Technology. RIPA buffer
was purchased from Santa Cruz Biotechnology, Inc. Clarity Western
ECL Substrate was from Bio-Rad. An apoptotic DNA-ladder kit was purchased
from Roche Applied Science.
3
Western Blot Analysis of HPV-18 Infection
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Frozen PHK and HPV-18 infected raft cultures were homogenized in mammalian cell lysis buffer and protein quantified as described [19 (link)]. Fifty or 100 µg of total protein were resolved in 5–15% gradient SDS-PAGE and wet transferred to PVDF membranes. Immunoblots were blocked in 5% nonfat dry milk or 5% BSA, probed with antibodies as described by the antibody suppliers. Signals were detected by GE Healthcare Amersham ECL Western Blotting Detection Reagents (Fisher Scientific, Waltham, MA, USA). The sources of various antibodies are as follows: HPV-18 E6, HPV-18 E7, (Santa Cruz (Dallas, TX, USA); p53 (DO7, Leica Biosystems-Novacastra Buffalo Grove, IL, USA); pRB, p130 (BD Biosciences. San Jose, CA, USA); total ATM, phospho-ATM (S1981; Abcam; Cambridge, MA, USA); ATR, total Chk1, ph-Chk1 (S296), ph-Chk1 (S345), Chk2, pChk2 (T68), γ-H2AX (S139) and cleaved PARP and cleaved caspase 3 (Cell Signaling Technologies; Danvers, MA, USA). Each experiment was repeated two or more times.
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