重组Anti-Lamin A + Lamin C抗体[EPR4100] -核Envelope Marker

• WB

Lab

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

ab108595 was shown to specifically react with Lamin A + C (LMNA) in wild type HAP1 cells. No band was observed when Lamin A + C (LMNA) knockout samples were used. Wild-type and Lamin A + C (LMNA) knockout samples were subjected to SDS-PAGE. The membrane was blocked for an hour using 5% milk before ab108595 and ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at a 1/10000 and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (ab108595) at 1/10000 dilution

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

Lamin A/C knockout HAP1 whole cell lysate at 20 µg

Lane 3:

HeLa whole cell lysate at 20 µg

Predicted band size: 74 kDa

Observed band size: 64 kDa,76 kDa

false

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemical analysis of formalin fixed paraffin embedded human liver labelling Lamin A + Lamin C with ab108595 at a concentration of 0.01µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was performed with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5 for 32 mins. ab108595 anti-Lamin A + Lamin C antibody [EPR4100] was incubated for 16 mins at 37°C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual).

• WB

Lab

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Western blot : Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker ab108595 staining at 1/10000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 70/75 kDa in Wild-type MCF7 cell lysates with no signal observed at this size in LMNA knockout MCF7 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5pc 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 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (ab108595) at 1/1000 dilution

Lane 1:

Wild-type MCF7 at 20 µg

Lane 2:

Western blot - Human LMNA knockout MCF7 cell line (ab287603) at 20 µg

Lane 3:

Wild-type HAP1 at 20 µg

Lane 4:

Knockout HAP1 at 20 µg

Lane 5:

HeLa at 20 µg

Secondary

Lanes 1 - 5:

Goat anti-Rabbit 800CW at 1/20000 dilution

Lanes 1 - 5:

Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 74 kDa

Observed band size: 70 kDa,75 kDa,37 kDa

false

• WB

Unknown

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

All lanes:

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (ab108595) at 1/10000 dilution

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma) cell lysates at 10 µg

Lane 2:

SH-SY5Y (Human neuroblastoma cell line from bone marrow) cell lysates at 10 µg

Predicted band size: 74 kDa

false

• WB

Lab

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

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

All lanes:

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (ab108595) at 1/100000 dilution

All lanes:

HeLa (Human epithelial cell line from cervix adenocarcinoma) cell lysate at 10 µg

Secondary

All lanes:

Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

Predicted band size: 74 kDa

Observed band size: 63 kDa,74 kDa

false

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colonic carcinoma tissue labeling Lamin A + C with ab108595.

Green - Lamin A + C.

Red - PI.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human liver tissue labeling Lamin A + C with ab108595 at 1/250 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human urothelial carcinoma tissue labeling Lamin A + C with ab108595.

Green - Lamin A + C.

Red - PI.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervix carcinoma tissue labeling Lamin A + C with purified ab108595 at 1/250. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Counterstained with hematoxylin.

Negative control using PBS instead of primary antibody (inset).

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Histological analyses of hiPSCs (Human induced pluripotent stem cell) transplanted into the subretinal space of nude rats.

Eye balls were excised from a nude rat 7 weeks after subretinal transplantation with 1×10⁴ hiPSCs. Transplanted tissues were fixed with 4% paraformaldehyde. Paraffin embedded tissue sections were stained with haematoxylin/eosin. Then, the paraffin sections were deparaffinized with xylene and sequential 100%, 95%, 80%, 70% ethanol treatments for 5 min each. The sections were treated with 10 mM citric acid (pH 6) at 95°C for 50 min followed by permeation with 0.4% Triton-X in PBS at room temperature for 30 min.

The deparaffinized sections were stained with ab108595 (Panel M), Hoechst 33258 (Panel N).

Kanemura et al PLoS One. 2014 Jan 14;9(1):e85336. doi: 10.1371/journal.pone.0085336. eCollection 2014. Fig 6. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue labeling Lamin A + C with ab108595.

Green - Lamin A + C.

Red - PI.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Intracellular Flow Cytometry analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Lamin A + C with purified ab108595 at 1/110 (red). Cells were fixed with 2% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/150) was used as the secondary antibody.

Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabeled control, cells without incubation with primary and secondary antibodies.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunocytochemistry/Immunofluorescence analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Lamin A + C (green) with purified ab108595 at 1/500. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain.

Control : Primary antibody (1/500) and secondary antibody ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/500).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunocytochemsitry/Immunofluorescence analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Lamin A + C with unpurified ab108595 at 1/250 dilution.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Overlay histogram showing HeLa (Human epithelial cell line from cervix adenocarcinoma) cells stained with unpurified ab108595 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab108595 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluorr^® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x10⁶ cells) used under the same conditions. Unlabeled sample (blue line) was also used as a control. Acquisition of >5000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human brain tissue labeling Lamin A + C with ab108595 at 1/250 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervical carcinoma tissue labeling Lamin A + C with ab108595.

Green - Lamin A + C.

Red - PI.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IP

Lab

Immunoprecipitation - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

ab108595 (purified) at 1/30 immunoprecipitating Lamin A + C in HeLa (Human epithelial cell line from cervix adenocarcinoma) cell lysate. For western blotting, a peroxidase-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/1000).

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

All lanes:

Immunoprecipitation - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (ab108595)

Predicted band size: 74 kDa

Observed band size: 74 kDa

false

• WB

Supplier Data

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

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

Western blot analysis of HEK-293T cells transfected with a human progerin expression vector containing a Flag-tag whole cell lysate 2 ug mixed with HeLa whole cell lysate 20 ug. The membrane was cut into strips and each strip was incubated separately with the following antibodies : Lane 1 : anti Lamin A + Lamin C antibody ab108595 (1 : 1000), Lane 2 : anti Flag-tag antibody (1 : 10000), Lane 3 : anti-progerin ab320642 (1 : 1000).

The anti-progerin antibody ab320642 specifically detects human progerin.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 12714972, PMID : 33408413, PMID : 34694158).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, Anti-Lamin A + Lamin C antibody - (ab108595) staining at 64-76KDa dilution.

In Western blot, Anti-Flag tag staining at 1/10000 dilution.

All lanes:

Western blot - Anti-Progerin antibody [EPR28694-72] (<a href='/products/primary-antibodies/progerin-antibody-epr28694-72-ab320642'>ab320642</a>) at 1/1000 dilution

All lanes:

HEK-293T (human embryonic kidney epithelial cell) cells transfected with a human progerin expression vector containing a Flag-tag whole cell lysate 2 μg mixed with HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 74 kDa,64-76 kDa

false

Exposure time: 81s

• WB

Lab

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

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

All lanes:

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (ab108595) at 1/100000 dilution

Lane 1:

HepG2 (Human liver hepatocellular carcinoma cell line) cell lysate at 20 µg

Lane 2:

HaCaT (Human keratinocyte cell line) cell lysate at 20 µg

Secondary

All lanes:

Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

Predicted band size: 74 kDa

Observed band size: 63 kDa,74 kDa

false

• WB

Supplier Data

Western blot - Anti-Lamin A + Lamin C antibody [EPR4100] - Nuclear Envelope Marker (AB108595)

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

This antibody does not cross-react with human Lamin A and Lamin C.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, Anti-Lamin A + Lamin C antibody - (ab108595) staining at 64-76KDa dilution.

In Western blot, Anti-Flag tag staining at 1/10000 dilution.

In Western blot, (Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-Progerin antibody [EPR28694-72] (<a href='/products/primary-antibodies/progerin-antibody-epr28694-72-ab320642'>ab320642</a>) at 1/1000 dilution

Lane 1:

HEK-293T (human embryonic kidney epithelial cell) cells transfected with an empty vector containing a DDDDK-tag, whole cell lysate at 20 µg

Lane 2:

HEK-293T cells transfected with a human progerin expression vector containing a Flag-tag, whole cell lysate at 20 µg

Lane 3:

HEK-293T cells transfected with a human Lamin C expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 4:

HEK-293T cells transfected with a human Lamin A expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 74 kDa,36 kDa,64-76 kDa

false

Exposure time: 10s