Anti activated caspase 3

Manufactured by Cell Signaling Technology

Sourced in United States

Anti-activated caspase-3 is a laboratory reagent used for the detection of activated caspase-3, a key executioner protein involved in apoptosis (programmed cell death). It is a specific antibody that binds to the cleaved (activated) form of caspase-3.

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Lab products found in correlation

Horseradish peroxidase conjugated anti rabbit igg, by Merck Group (2 mentions) Anti activated caspase 8, by Cell Signaling Technology (2 mentions) Anti rabbit igg, by Merck Group (2 mentions) Anti goat igg, by Santa Cruz Biotechnology (2 mentions) Anti β actin, by Merck Group (2 mentions) Anti actin, by Merck Group (2 mentions) Anti gfp, by Thermo Fisher Scientific (2 mentions) Anti caspase 3, by Cell Signaling Technology (2 mentions) Paraformaldehyde (pfa), by Merck Group (2 mentions) Anti nqo1, by Cell Signaling Technology (1 mentions) Anti c fos, by Santa Cruz Biotechnology (1 mentions) Target retrieval solution s1700, by Agilent Technologies (1 mentions) Histofine simple stainmax po secondary antibody system, by Nichirei Biosciences (1 mentions) Alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Alexa fluor 594, by Thermo Fisher Scientific (1 mentions) Anti ki67, by BD (1 mentions) Anti bax, by Cell Signaling Technology (1 mentions) Anti bcl 2, by Cell Signaling Technology (1 mentions) Anti cyclin b1, by Cell Signaling Technology (1 mentions) Anti total p53, by Cell Signaling Technology (1 mentions)

19 protocols using anti activated caspase 3

Immunohistochemical Analysis of Olfactory Tissue

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Mice were perfused with 4% paraformaldehyde in 0.1% phosphate buffer and post-fixed for 24 h in the same fixative. The head tissue, including the cochlea, OE, and olfactory bulb (OB), was decalcified with 10% EDTA solution, pH 7.0, and embedded in paraffin. Coronal sections were cut at 4 μm thickness and mounted on silane-coated slides. Deparaffinized sections were autoclaved at 121 °C for 20 min in Target Retrieval Solution (S1700; Dako) for antigen retrieval. Immunohistochemistry was performed with the following antibodies: anti-olfactory marker protein (OMP, goat polyclonal, 1:2000 dilution; Wako Chemicals), anti-NQO1 (rabbit polyclonal, 1:300; Cell Signaling Technology), anti-activated caspase-3 (rabbit polyclonal, 1:300; Cell Signaling Technology), anti-Ki67 (mouse monoclonal, 1:300; BD Biosciences), anti-c-fos (rabbit polyclonal, 1:50; Santa Cruz Biotechnology), anti-MnSOD (rabbit monoclonal, 1:100; Epitomics Inc.), and anti-8-hydroxy-2′-deoxyguanosine (8-OHdG, goat polyclonal antibody, 1:100; Alpha Diagnostic International Inc.). Immunoreactivity was detected using the following antibodies in the Histofine Simple StainMAX-PO secondary antibody system (Nichirei) according to the manufacturers’ instructions, donkey anti-goat Alexa Fluor 488 (1:100; Invitrogen), and donkey anti-rabbit Alexa Fluor 594 (1:100; Invitrogen), incubated for 1 h at room temperature.

Tuerdi A., Kikuta S., Kinoshita M., Kamogashira T., Kondo K., Iwasaki S, & Yamasoba T. (2018). Dorsal-zone-specific reduction of sensory neuron density in the olfactory epithelium following long-term exercise or caloric restriction. Scientific Reports, 8, 17300.

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Comprehensive Immunoblotting Approach

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Western immunoblots were run as described previously 21 (link). Primary antibodies and their sources were as follows. Anti-total p53, anti-cyclin B1, anti-Bax, anti-Bcl-2, anti-CDK1, anti-phospho-CDK1, anti-activated caspase 8, anti-activated caspase 9, and anti-activated caspase 3 were from Cell Signaling Technology. Anti-ZIKV Env monoclonal antibody was purchased from Zoongen Co., Ltd (Beijing, China). Anti-p21^Cip1/Waf1 and anti-6×His tag was from Abcam. Anti-β-actin and the secondary antibodies horseradish peroxidase-conjugated anti-rabbit IgG and anti-rabbit IgG were from Sigma-Aldrich. Anti-goat IgG was from Santa Cruz Biotechnology.

Liu J., Li Q., Li X., Qiu Z., Li A., Liang W., Chen H., Cai X., Chen X., Duan X., Li J., Wu W., Xu M., Mao Y., Chen H., Li J., Gu W, & Li H. (2018). Zika Virus Envelope Protein induces G2/M Cell Cycle Arrest and Apoptosis via an Intrinsic Cell Death Signaling Pathway in Neuroendocrine PC12 Cells. International Journal of Biological Sciences, 14(9), 1099-1108.

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Quantifying Otic Cell Proliferation and Apoptosis

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Embryos were fixed in 4% paraformaldehyde, 0.1% Tween-20 overnight; washed with phosphate-buffered saline, 0.1% Tween-20; embedded in 3.5% low melting point agarose; and cut into 40 μm-thick transverse sections by vibratome. Sections were processed for immunohistochemistry as described (Shim et al., 2005 (link)). Primary antibodies were: anti-phospho-histone-H3 (Millipore 05-806, 1:1,000) and anti-activated-caspase-3 (Cell Signaling 9661, 1:200). Secondary antibodies were labeled with either Alexa-488 or Alexa-568 (Life Technologies, 1:1,000). DAPI (Sigma) was used to counter-stain the nuclei.
Vibratome sections containing the otic placode were imaged using a Zeiss LSM510 laser scanning confocal microscope. In each vibratome section, the total number of otic cells from both placodes, staining for either phospho-histone-H3 or activated caspase-3 was counted. The volume of the otic tissue in each vibratome section was calculated as the area of the otic region in the central image from the confocal z-stack (mm², measured in ImageJ), multiplied by 0.04 mm (the thickness of each vibratome section). For each embryo, the total number of pH3+ or activated caspase-3+ cells was normalized by the total volume of otic tissue measured (number/mm³). Significance of differences between genotypes was measured by ANOVA.

Zhang J., Wright K.D., Mahoney Rogers A.A., Barrett M.M, & Shim K. (2014). Compensatory regulation of the size of the inner ear in response to excess induction of otic progenitors by FGF signaling. Developmental dynamics : an official publication of the American Association of Anatomists, 243(10), 1317-1327.

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Apoptotic Signaling Pathways Evaluation

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DMSO, transresveratrol (RES), curcumin from Curcuma Longa (CUR), Sulforhodamine B (SRB), staurosporine and Hoechst 33342 were purchased from Sigma-Aldrich (Milan, Italy). Rabbit polyclonal anti-Bax and mouse monoclonal anti-Bcl-2 antibodies were obtained from BD Pharmingen (BD Biosciences, San Jose, CA, USA). Antibodies against PARP-1, ERK1/2 (C-14), phospho-ERK (E-4), NF-κB and p53 (DO-1) were obtained from Santa Cruz Biotechnology (CA, USA). Rabbit polyclonal anti-actin was purchased from Sigma-Aldrich (Milan, Italy). The anti-activated caspase 3, anti-caspase 9, anti-caspase 8, anti-AKT and anti-phospho-AKT antibodies were purchased from Cell Signaling Technology (MA, USA). Antibody against LC3 was obtained from Novus Biologicals (Littleton, CO, USA). Goat anti-mouse IgG Alexa fluor-488-conjugated antibody was purchased from Life Technologies™ Molecular Probes (Oregon, USA). The goat anti-mouse or -rabbit IgG peroxidase conjugated secondary antibodies were obtained from Sigma-Aldrich (Milan, Italy).

Masuelli L., Stefano E.D., Fantini M., Mattera R., Benvenuto M., Marzocchella L., Sacchetti P., Focaccetti C., Bernardini R., Tresoldi I., Izzi V., Mattei M., Frajese G.V., Lista F., Modesti A, & Bei R. (2014). Resveratrol potentiates the in vitro and in vivo anti-tumoral effects of curcumin in head and neck carcinomas. Oncotarget, 5(21), 10745-10762.

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Epidermal Dissection and Immunostaining

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Dissection and immunostaining of larval epidermis were performed as previously described.^{52 (link)} Primary antibodies: anti-Fasciclin III (Developmental Studies Hybridoma Bank, Iowa City, IA, USA, 1:50), anti-activated Caspase-3 (Cell Signaling, Danvers, MA, USA, 1:150), and anti-GFP (Life Technologies, Basel, Switzerland, 1:500). Secondary antibodies: alexa488-conjugated anti-mouse IgG (Life Technologies, 1:1000), Cy3-conjugated goat anti-rabbit IgG (Jackson ImmunoResearch, West Grove, PA, USA, 1:1000). TUNEL labeling kit (Roche, Basel, Switzerland) was used to label apoptotic cells.

Jo J., Im S.H., Babcock D.T., Iyer S.C., Gunawan F., Cox D.N, & Galko M.J. (2017). Drosophila caspase activity is required independently of apoptosis to produce active TNF/Eiger during nociceptive sensitization. Cell Death & Disease, 8(5), e2786-.

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Immunohistochemical Analysis of Mouse Brain

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The anesthetized mice were perfused with with 4% paraformaldehyde (PFA) dissolved in phosphate buffered saline (PBS), and isolated brains were post-fixed with 4% PFA at 4°C for over night. After washing with PBS, the brains were cryoprotected with 30% sucrose solution, and embedded in Tissue-Tek. The frozen brains were sectioned at a thickness of 35 μm using a Cryostat (CM1900 and CM1850, Leica), and incubated with primary antibodies, including anti-GFP (rabbit polyclonal, Life Technology or rat monoclonal, Nakarai Tesque), anti-RFP (rabbit polyclonal, Abcam), anti-GLAST (goat polyclonal, Frontier Institute), anti-GFAP (mouse monoclonal, SIGMA), anti-ß-gal (rabbit polyclonal, MP Biomedicals), anti-activated caspase3 (rabbit polyclonal, Cell signaling), and anti-S100 (rabbit polyclonal, DAKO). After washing, the sections were incubated with secondary antibodies, including Alexa-Fluor 488, 594 or 633-conjugated anti-rabbit, anti-mouse, anti-rat and anti-goat antibodies (Life technologies). Nuclear staining was performed with Hoechst 33258. Sections were analyzed with either a fluorescent microscope (BX51, OLYMPUS) equipped with a cooled CCD camera system (DP71, OLYMPUS) or a laser confocal microscope (FV1000D, OLYMPUS).

Nomura T., Nishimura Y., Gotoh H, & Ono K. (2016). Rapid and efficient gene delivery into the adult mouse brain via focal electroporation. Scientific Reports, 6, 29817.

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Immunohistochemical Profiling of Neurological Markers

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Antibodies used in this study are: anti-pan-flavivirus (MAB10216, clone D1-4G2) anti NeuN (Abcam), anti Iba-1 (Abcam), anti Tau (Abcam), anti-GFAP (Abcam), anti-caspase 3 and anti-activated caspase 3 (Cell Signalling Technology) and secondary antibodies coupled to Alexa dyes (488, 555 or 647, Thermofischer Scientific).

Salinas S., Constant O., Desmetz C., Barthelemy J., Lemaitre J.M., Milhavet O., Nagot N., Foulongne V., Perrin F.E., Saiz J.C., Lecollinet S., Van de Perre P, & Simonin Y. (2017). Deleterious effect of Usutu virus on human neural cells. PLoS Neglected Tropical Diseases, 11(9), e0005913.

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Drosophila Larval Epidermis Dissection

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The third instar larval epidermis was dissected and processed as detailed previously (Burra et al., 2013 (link)). To highlight wound morphology, a mouse monoclonal antibody against Fasciclin III was used (1:50; Developmental Studies Hybridoma Bank). Rat anti-Sd was a gift from Kirsten Guss (Guss et al., 2013 (link)) and was used at 1:20. Mouse anti-GFP monoclonal antibody (Life Technologies) was used at 1: 500. Mouse anti-β-galactosidase monoclonal antibody (Promega #Z3781) was used at 1:1000. Anti-activated Caspase 3 (Cell Signaling) was used at 1:150. Mouse anti-Drosophila Profilin (Developmental Studies Hybridoma Bank) was used undiluted. Rabbit anti-DsRed (Clontech) was used at 1:1000. Rabbit anti-Lethal (2) giant larvae (l(2)gl) was used at 1:50 (Santa Cruz, sc-98260). Alexa 546 conjugated Phalloidin (Invitrogen) was used at 1:200. A TUNEL staining kit (Roche Diagnostics) was also used to detect DNA double-stranded breaks.

Tsai C.R., Anderson A.E., Burra S., Jo J, & Galko M.J. (2017). Yorkie regulates epidermal wound healing in Drosophila larvae independently of cell proliferation and apoptosis. Developmental biology, 427(1), 61-71.

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Investigating HIF-2α Inhibition and Autophagy

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Hydroxychloroquine, temsirolimus (CCI-779), and 5-fluorouracil (5-FU) were purchased from Sigma-Aldrich (St. Louis, MO, USA). The HIF-2α-specific inhibitor PT-2385 was purchased from Biovision. Dihydroethidium (DHE) was obtained from Invitrogen^TM (Waltham, MA, USA) (Cat. No. D11347). N-Acetyl-L-cystein (NAC) was obtained from Sigma-Aldrich (Cat. No. A7250). The antibodies used in the experiments were from the following sources: anti-LC3 from Novus Biologicals (Centennial, CO, USA); and anti-activated caspase-3, anti-HIF-1α, anti-HIF-2α, anti-SQSTM1/p62, anti-β-tubulin, anti-Atg7, anti-Phospho-S6 Ribosomal protein (Ser235/236), anti-Beclin.1 and anti-PI3K-Class III were all obtained from Cell Signaling Technology (Danvers, MA, USA). Goat anti-mouse and anti-rabbit IgG-horseradish peroxidase-conjugates were from Pierce (Rockford, IL, USA)

Saint-Martin A., Martínez-Ríos J., Castañeda-Patlán M.C., Sarabia-Sánchez M.A., Tejeda-Muñoz N., Chinney-Herrera A., Soldevila G., Benelli R., Santoyo-Ramos P., Poggi A, & Robles-Flores M. (2019). Functional Interaction of Hypoxia-Inducible Factor 2-Alpha and Autophagy Mediates Drug Resistance in Colon Cancer Cells. Cancers, 11(6), 755.

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Protein Expression Analysis by Western Blot

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Western blot analysis was performed as described previously [16 (link)]. The primary antibodies were the following: anti-E-cadherin (1:1000, Cell Signaling Technology, USA), anti-N-cadherin (1:1000, Cell Signaling Technology), anti-Vimentin (1:1000, Abcam, USA), anti-activated caspase-3 (1:1000, Cell Signaling Technology), anti-total caspase-3 (1:1000, Cell Signaling Technology), anti-c-Met (1:1000, Abcam), and anti-GAPDH (1:1000, Santa Cruz Biotechnology, USA).

Liu F., Chen N., Gong Y., Xiao R., Wang W, & Pan Z. (2017). The long non-coding RNA NEAT1 enhances epithelial-to-mesenchymal transition and chemoresistance via the miR-34a/c-Met axis in renal cell carcinoma. Oncotarget, 8(38), 62927-62938.

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