Anti-Cleaved Caspase-3 抗体 [E83-77]

• WB

Lab

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (AB32042)

Lane 1 : Wild type HAP1 + DMSO for 24 hours, whole cell lysate (20 μg)
Lane 2 : Wild type HAP1 + 2uM Staurosporine (ab146588) for 24 hours, whole cell lysate (20 μg)
Lane 3 : HAP1 CASP3 KO + DMSO for 24 hours, whole cell lysate (20 μg)
Lane 4 : HAP1 CASP3 KO + 2uM Staurosporine (ab146588) for 24 hours, whole cell lysate (20 μg)
Lane 5 : HeLa + DMSO for 24 hours, whole cell lysate (20 μg)
Lane 6 : HeLa + 2uM Staurosporine (ab146588) for 24 hours, whole cell lysate (20 μg)
Lanes 1 - 6 : Merged signal (red and green). Green - ab32042 observed at 17 kDa. Red - loading control, ab130007, observed at 130 kDa.

ab32042 was shown to specifically react with CASP3 (Caspase-3) when CASP3 (Caspase-3) knockout samples were used. HAP1 wild-type and CASP3 (Caspase-3) knockout samples were subjected to SDS-PAGE. ab32042 and ab130007 (Mouse anti vinculin loading control) were incubated overnight at 4°C at 500 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

Cells were grown to confluency prior to treatment.

All lanes:

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042)

Predicted band size: 32 kDa

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• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-Cleaved Caspase-3 antibody [E83-77] (AB32042)

High-glucose induces apoptosis in human Vascular endothelial cells (VECs).

Apoptotic responses in VEC were analyzed by detection of cleaved-Caspase-3 immunofluorescence using ab32042. Cells were treated with low glucose (LG) or high glucose (HG) for 72 hours before treated with 100 ng/mL bFGF, 1 μM sp600125 (sp), or 1 μM U0126 (U) or 10 μM MnTmPyP for 1 hour. Bar = 100 μm.

Image from Zhu ZX et al., PLoS One. 2015;10(12):e0144495. Fig 7.; doi: 10.1371/journal.pone.0144495. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

• WB

Supplier Data

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (AB32042)

Western blot : Anti-CASP3 antibody [E83-77] (ab32042) staining at 1/500 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab32042 was shown to bind specifically to CASP3. A band was observed at 16/28 kDa in treated wild-type HeLa cell lysates with no signal observed at this size in CASP3 knockout cell line. To generate this image, wild-type and CASP3 knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042) at 1/500 dilution

Lane 1:

Wild-type HeLa Treated Staurosporine (2uM, 4h) cell lysate at 20 µg

Lane 2:

CASP3 knockout HeLa Treated Staurosporine (2uM, 4h) cell lysate at 20 µg

Lane 3:

Wild-type HeLa Vehicle Control Staurosporine (0uM, 4h) cell lysate, at 20 µg

Lane 4:

CASP3 knockout HeLa Vehicle Control Staurosporine (0uM, 4h) cell lysate at 20 µg

Predicted band size: 32 kDa

Observed band size: 16 kDa,28 kDa

false

• WB

Unknown

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (AB32042)

Lanes 1 - 2:

anti Pro Caspase 3 at 1/10000 dilution

Lanes 3 - 4:

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042) at 1/500 dilution

Lane 1:

Jurkat (human T cell leukemia cell line from peripheral blood) cell lysate

Lanes 2 and 4:

Jurkat cell lysate + Camptothecin

Lane 3:

Jurkat cell lysate

Predicted band size: 32 kDa

Observed band size: 17 kDa,30 kDa

false

• WB

Ap

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (AB32042)

All lanes:

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042) at 1/500 dilution

Lane 1:

HeLa Whole Cell Lysate (2 uM Staurosporine, 4Hr) at 20 µg

Lane 2:

HeLa Whole Cell Lysate (untreated) at 20 µg

Lane 3:

Cleaved Caspase 3 (recombinant protein) at 0.1 µg

Secondary

All lanes:

800CW Goat Anti-Rabbit IgG at 1/10000 dilution

Predicted band size: 32 kDa

Observed band size: 17 kDa

false

• ICC/IF

CiteAb

Immunocytochemistry/ Immunofluorescence - Anti-Cleaved Caspase-3 antibody [E83-77] (AB32042)

Immunocytochemistry-immunofluorescence using Anti-Cleaved Caspase-3 antibody [E83-77], ab32042. Publication image from Capogrossi, M. C. et al., 2010, Mol Cancer, 20412563. Legend direct from paper.

RGDS treatment activates caspase-3. A : SK-MEL-110 seeded on collagen-IV were incubated for 48 h with FGF-2 in the presence or in the absence of RGDS or RGES (500 µg/ml). Top : Pro-caspase 3 cleavage and caspase-3 activation were observed by western blotting. Bottom : caspase-3 activation was confirmed by confocal microscopy using an anti-active caspase-3 antibody. B : Pre-treatment (2 h) of melanoma cells with Z-VAD-FMK (50 µM), a general caspases inhibitor, abolished the RGDS anti-proliferative effect (p < 0.005 vs FGF-2 in the presence of Z-VAD-FMK). All these experiments were carried out three times in duplicate.

• WB

CiteAb

Western blot - Anti-Cleaved Caspase-3 antibody [E83-77] (AB32042)

Western Blotting using Anti-Cleaved Caspase-3 antibody [E83-77], ab32042. Publication image from Chen, W. et al., 2019, Nat Commun, 31792210. Legend direct from paper.

ELP5 promotes P53 expression by IRES-dependent translation. a ELP5 depletion substantially rescued the accumulation and activation of P53 and P53-mediated apoptosis under GEM treatment at IC50 for 72 h in GBC cells. b RT-qPCR confirmed that ELP5 depletion did not affect the P53 mRNA level with or without GEM treatment, but P53 target genes (i.e., P21 and MDM2) displayed dramatically reduced mRNA levels in ELP5−/− cells. c ELP5 depletion could not affect the P53 degradation rate in GBC cells. The P53 protein level was normalized to that of β-Actin, and the P53 protein levels in each cell were set as 1 at the 0 time point. d Schematic drawing of the exogenously delivered P53 construct with or without an IRES sequence within the 5′-UTR of the P53 ORF sequence, IRES-P53-Flag, and P53-Flag, respectively. UTR, untranslated region; ORF, open-reading frame. e The IRES-P53-Flag construct was inefficiently expressed in ELP5-depleted cells, but P53-Flag construct was adequately expressed in ELP5-depeleted and WT cells. f Schematic drawing of the bicistronic construct inserted with P53 IRES between Rluc and Fluc. Rluc, Renilla luciferase; Fluc, Firefly luciferase. gP53 IRES activity was notably reduced in ELP5-depleted cells, as assessed by the ratio of Fluc to Rluc via dual-luciferase reporter assay. h, i Ectopically expressed ELP5 in P53-knockout cells (h) could not rescue the reduced GEM-induced apoptosis caused by P53 depletion (i). Data represent the mean ± S.D., n = 3 independent experiments in b, c, g, i, error bars represent S.D. Unpaired Student’s t tests were used in b, g, i (NS, non-significant, *P < 0.05, **P < 0.01 and ***P < 0.001).

false