Anti-Hsp27 抗体 [G3.1]

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Hsp27 antibody [G3.1] (AB2790)

Flow cytometry overlay histogram of [4% Paraformaldehyde] HeLa (Human cervix adenocarcinoma epithelial cell) permeabilised with 90% methanol labelling Hsp27 with ab2790 at 1/1000 dilution (Red) compared with a mouse monoclonal IgG isotype control (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Secondary antibody details Goat anti mouse IgG (Alexa Fluor^® 488) (ab150113), at 1/2000 dilution.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Hsp27 antibody [G3.1] (AB2790)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa cells labelling Hsp27 with ab2790 at 1/100 dilution, followed by AlexaFluor® 488-conjugated Goat anti-Mouse secondary antibody (ab150113) at 1/1000 dilution at RT for 45 min. ab179513 Anti-beta Tubulin, used as a counterstain at 1/200 dilution, was co-incubated with ab9509 overnight at 4° C, followed by Alexa Fluor® 594 Goat Anti-Rabbit secondary (ab150080) at 1/1000 dilution at RT for 45 min. Nucleus were visualized using DAPI.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hsp27 antibody [G3.1] (AB2790)

Immunohistochemical analysis of paraffin-embedded human tissues labeling Hsp27 with ab2790 at 1/1000 dilution followed by Goat Anti-Mouse IgG H&L (HRP polymer) (ab214879). The section was incubated with ab2790 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with hematoxylin.

• IP

Lab

Immunoprecipitation - Anti-Hsp27 antibody [G3.1] (AB2790)

Hsp27 was immunoprecipitated from HeLa (human cervix adenocarcinoma epithelial cell), whole cell lysate 10 μg with ab2790 1/1000 dilution (0.45 μg/ml). Western blot was performed on the immunoprecipitate using ab2790 at 1/1000 dilution. Capture antibody, 1/30 dilution (2μg in 0.35mg lysates).

Mouse monoclonal IgG1, kappa (ab18443) Isotype Control was used instead of ab2790 in HeLa whole cell lysate (Lane 3).

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Secondary antibody (ab131368), mouse IgG for IP (HRP) used at 1/1000 dilution.

All lanes:

Immunoprecipitation - Anti-Hsp27 antibody [G3.1] (ab2790) at 1/1000 dilution

All lanes:

HeLa (human cervix adenocarcinoma epithelial cell ) whole cell lysate at 10 µg

Secondary

All lanes:

Immunoprecipitation - Anti-mouse IgG for IP (HRP) (<a href='/products/secondary-antibodies/mouse-igg-for-ip-hrp-ab131368'>ab131368</a>) at 1/1000 dilution

Observed band size: 27 kDa

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Exposure time: 7s

• WB

Lab

Western blot - Anti-Hsp27 antibody [G3.1] (AB2790)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Hsp27 antibody [G3.1] (ab2790) at 1/1000 dilution

Lane 1:

Human liver tissue lysate at 20 µg

Lane 2:

Human heart tissue lysate at 20 µg

Lane 3:

HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

549 (Human lung carcinoma epithelial cell) whole cell lysate at 20 µg

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• WB

CiteAb

Western blot - Anti-Hsp27 antibody [G3.1] (AB2790)

Western Blotting using Anti-Hsp27 antibody [G3.1], ab2790. Publication image from Kim, M. J. et al., 2023, Nat Commun, 37433777. Legend direct from paper.

Small HSP and HSP110 families have a tendency to be increased under mitochondrial stress.a RNA-seq analysis of HSPs gene expression log2 fold changes (log2FC) in NDUFA11 KO and NDUFA13 KO compared to WT HEK293T cells (n = 4). Up- and down-regulated genes (q-value < 0.05) are shown in green and pink, respectively. The intensity of the color shades depends on the level of expression change. Gray indicates genes with not statistically significant expression changes. b mRNA expression patterns of selected transcripts validated by RT-qPCR. The mRNA levels are presented as fold changes relative to WT. Data shown are mean ± SD (n = 3 biological replicates with two technical replicates). p-value from an ordinary one-way ANOVA with Dunnett’s multiple comparisons test using GraphPad Prism. c Western blot analysis of HSPs expression performed in whole cell lysates of NDUFA11 KO, NDUFA13 KO and WT HEK293T cells. ACTB was used as a loading control. Data shown are representative of three independent experiments. d Quantification of HSPs in western blot analysis normalized to ACTB using ImageJ. The protein levels are presented as fold changes relative to WT. Data shown are mean ± SD (n = 3). p-value from two-sided, unpaired t-test using GraphPad Prism. Source data are provided as a Source Data file.

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