Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

This data was developed using ab229025 the same antibody clone in a different buffer formulation.

ab229025 staining LMNB1 in Hap1 WT and Hap1 LMNB1 KO cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab229025 at 0.2µg/ml (shown in green) and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control (shown in pseudocolour magenta). Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor^® 488), pre-adsorbed at 1/1000 dilution. Nuclear DNA was labelled with DAPI (shown in blue).

Also suitable in cells fixed with 100% methanol (5 min).

Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a single confocal section is shown.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Immunohistochemical analysis of paraffin-embedded human breast tissue stained for Lamin B1 using ab229025 at 1/2000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Nuclear envelope staining on human breast (PMID : 26469707) is observed. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (a229025).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling Lamin B1 (Green) with ab229025 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution. Confocal image showing nuclear membranous staining in HeLa cells. Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) was used as a counterstain at 1/200 dilution. The nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Alexa Fluor^®488 Goat anti-Rabbit secondary (ab150077) at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cell line labeling Lamin B1 with ab229025 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG isotype control (ab172730) (Black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/2000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Immunohistochemical analysis of paraffin-embedded human endometrial carcinoma stained for Lamin B1 using ab229025 at 1/2000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer). Nuclear envelope staining on human endometrial carcinoma is observed. Counterstained with hematoxylin. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). The section was incubated with ab229025 at 4°C overnight This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

This data was developed using ab229025 the same antibody clone in a different buffer formulation.

ab229025 staining LMNB1 in HeLa WT and HeLa LMNB1 KO cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab229025 at 0.2µg/ml (shown in green) and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control (shown in pseudocolour magenta). Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor^® 488), pre-adsorbed at 1/1000 dilution. Nuclear DNA was labelled with DAPI (shown in blue).

Also suitable in cells fixed with 100% methanol (5 min).

Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a single confocal section is shown.

• IP

Supplier Data

Immunoprecipitation - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Lamin B1 was immunoprecipitated from 0.35 mg HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab229025 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab229025 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

Lane 1 : HeLa whole cell lysate 10 μg (input).
Lane 2 : ab229025 IP in HeLa whole cell lysate (+).
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab229025 in HeLa whole cell lysate.

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

A very weak band around 66 kDa detected in lane 3 is due to spill over from lane2 (+).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

All lanes:

Immunoprecipitation - Anti-Lamin B1 antibody [EPR22165-121] - Nuclear Envelope Marker (<a href='/en-us/products/primary-antibodies/lamin-b1-antibody-epr22165-121-nuclear-envelope-marker-ab229025'>ab229025</a>)

Predicted band size: 66 kDa

Observed band size: 66 kDa

false

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Immunohistochemical analysis of paraffin-embedded mouse liver tissue stained for Lamin B1 using ab229025 at 1/2000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Nuclear envelope staining on mouse liver (PMID : 19925772) is observed. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Immunohistochemical analysis of paraffin-embedded rat cerebrum tissue stained for Lamin B1 using ab229025 at 1/2000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Nuclear envelope staining on rat cerebrum (PMID : 26469707, PMID : 19925772) is observed. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryonic fibroblast cell line) cells labeling Lamin B1 (Green) with ab229025 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution. Confocal image showing nuclear membranous staining in NIH/3T3 cells. Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) was used as a counterstain at 1/200 dilution. The nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Alexa Fluor^®488 Goat anti-Rabbit secondary (ab150077) at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized NIH/3T3 (mouse embryo fibroblast cell line) cell line labeling Lamin B1 with ab229025 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG isotype control (ab172730) (Black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/2000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

• IP

Supplier Data

Immunoprecipitation - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

Lamin B1 was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate with ab229025 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab229025 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

Lane 1 : NIH/3T3 whole cell lysate 10 μg (input).
Lane 2 : ab229025 IP in NIH/3T3 whole cell lysate (+).
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab229025 in NIH/3T3 whole cell lysate.

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

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

All lanes:

Immunoprecipitation - Anti-Lamin B1 antibody [EPR22165-121] - Nuclear Envelope Marker (<a href='/en-us/products/primary-antibodies/lamin-b1-antibody-epr22165-121-nuclear-envelope-marker-ab229025'>ab229025</a>)

Predicted band size: 66 kDa

Observed band size: 66 kDa

false

• WB

Lab

Western blot - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

This data was developed using the same antibody clone in a different buffer formulation (ab229025).

Lanes 1- 2 : Merged signal (red and green). Green - ab229025 observed at 66-70 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

ab229025 was shown to react with lamin B1 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255404 (knockout cell lysate ab263825) was used. Wild-type HeLa and LMNB1 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab229025 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 dilution and a 1 in 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 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Lamin B1 antibody [EPR22165-121] - Nuclear Envelope Marker (<a href='/en-us/products/primary-antibodies/lamin-b1-antibody-epr22165-121-nuclear-envelope-marker-ab229025'>ab229025</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

LMNB1 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human LMNB1 (Lamin B1) knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-lmnb1-lamin-b1-knockout-hela-cell-line-ab255404'>ab255404</a>)

Predicted band size: 66 kDa

Observed band size: 66-70 kDa

false

• WB

Lab

Western blot - Anti-Lamin B1 antibody [EPR22165-121] - BSA and Azide free (AB239399)

ab229025 was shown to specifically react with Lamin B1 in wild-type HAP1 cells as signal was lost in Lamin B1 knockout cells. Wild-type and Lamin B1 knockout samples were subjected to SDS-PAGE. ab229025 and ab181602 (Rabbit anti-GAPDH loading control) were incubated 1 hour at room temperature at 1/1000 dilution and 1/200,000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100,000 dilution for 1 hour at room temperature before imaging. The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument using the ECL technique.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab229025).

All lanes:

Western blot - Anti-Lamin B1 antibody [EPR22165-121] - Nuclear Envelope Marker (<a href='/en-us/products/primary-antibodies/lamin-b1-antibody-epr22165-121-nuclear-envelope-marker-ab229025'>ab229025</a>) at 1/5000 dilution

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

Lamin B1 knockout HAP1 whole cell lysate at 20 µg

Lane 3:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate at 20 µg

Predicted band size: 66 kDa

Observed band size: 70 kDa

false

Exposure time: 8s