Anti-Lin28A antibody [EPR24286-100]

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemistry analysis of Formalin/PFA-fixed paraffin-embedded sections Rat testis tissue labelling Lin28A with ab279647 at 1 : 1000 dilution (B), Wilms Tumor Protein with ab267377 at 1 : 500 dilution (C) and DKKL1 with ab316308 at 1 : 2000 dilution (D). Opal Polymer HRP Ms + Rb was used as a secondary antibody, and DAPI was used for a nuclear counter stain. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

Panel A : merged staining of anti-Lin28A (green; Opal™520), anti-Wilms Tumor Protein (magenta; Opal™690) and anti-DKKL1 (gray; Opal™570) on rat testis.

Panel B : anti-Lin28A staining undifferentiated spermatogonia in rat testis.

Panel C : ant-Wilms Tumor Protein staining Sertoli cells in rat testis.

Panel D : ant-DKKL11 staining mature sperm cells in rat testis.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab279647, ab267377 and ab316308 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemistry analysis of Formalin/PFA-fixed paraffin-embedded sections Mouse testis tissue labelling Lin28A with ab279647 at 1 : 1000 dilution (B), STAT4 with ab284408 at 1 : 500 dilution (C) and SCP1 with [ab303520 at 1 : 500 dilution (D). Opal Polymer HRP Ms + Rb was used as a secondary antibody, and DAPI was used for a nuclear counter stain. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

Panel A : merged staining of anti-Lin28A (green; Opal™520), anti-STAT4 (magenta; Opal™690) and anti-SCP1 (gray; Opal™570) on mouse testis.

Panel B : anti-Lin28A staining undifferentiated spermatogonia in mouse testis.

Panel C : ant-STAT4 staining round spermatids (rs) and elongated spermatids (es) in mouse testis.

Panel D : ant-SCP1 staining spermatocytes in mouse testis.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab279647, ab284408 and ab303520 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemistry analysis of Formalin/PFA-fixed paraffin-embedded sections Rat testis labelling Lin28A with ab279647 at 1 : 1000 dilution (B), STAT4 with ab284408 at 1 : 500 dilution (C) and SCP1 with ab303520 at 1 : 500 dilution (D). Opal Polymer HRP Ms + Rb was used as a secondary antibody, and DAPI was used for a nuclear counter stain. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

Panel A : merged staining of anti-Lin28A (green; Opal™520), anti-STAT4 (magenta; Opal™690) and anti-SCP1 (gray; Opal™570) on rat testis.

Panel B : anti-Lin28A staining undifferentiated spermatogonia in rat testis.

Panel C : ant-STAT4 staining round spermatids (rs) and elongated spermatids (es) in rat testis.

Panel D : ant-SCP1 staining spermatocytes in rat testis.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab279647, ab284408 and ab303520 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemistry analysis of Formalin/PFA-fixed paraffin-embedded sections Mouse testis tissue labelling Lin28A with ab279647 at 1 : 1000 dilution (B), Wilms Tumor Protein with ab267377 at 1 : 500 dilution (C) and DKKL1 with ab316308 at 1 : 2000 dilution (D). Opal Polymer HRP Ms + Rb was used as a secondary antibody, and DAPI was used for a nuclear counter stain. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

Panel A : merged staining of anti-Lin28A (green; Opal™520), anti-Wilms Tumor Protein (magenta; Opal™690) and anti-DKKL1 (gray; Opal™570) on rat testis.

Panel B : anti-Lin28A staining undifferentiated spermatogonia in rat testis.

Panel C : ant-Wilms Tumor Protein staining Sertoli cells in rat testis.

Panel D : ant-DKKL11 staining mature sperm cells in rat testis.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab279647, ab267377 and ab316308 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HEK-293T (human embryonic kidney epithelial cell, Left panel) / NCCIT (human pluripotent embryonic carcinoma epithelial cell, Right panel) cells labeling Lin28A with ab279647 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

Negative control : HEK-293T (PMID : 25479749).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemical analysis of paraffin-embedded rat testis tissue labeling Lin28A with ab279647 at 1/1000 dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining on rat undifferentiated spermatogonia. The section was incubated with ab279647 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized NIH/3T3 (mouse embryonic fibroblast, Left panel) / F9 (mouse embryonal carcinoma epithelial cell, Right panel) cells labeling Lin28A with ab279647 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

Negative control : NIH/3T3 (PMID : 21962509).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized F9 (mouse embryonal carcinoma epithelial cell line) cells labelling Lin28A with ab279647 at 1/500 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing nucleolar and cytoplasmic staining in F9 cell line. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was DAPI (Blue).

Negative control : NIH/3T3 (PMID : 21962509).

Secondary antibody only control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemical analysis of paraffin-embedded mouse testis tissue labeling Lin28A with ab279647 at 1/1000 dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining on mouse undifferentiated spermatogonia (PMID : 32094118). The section was incubated with ab279647 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BON^D® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemical analysis of paraffin-embedded mouse liver tissue labeling Lin28A with ab279647 at 1/1000 dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). The section was incubated with ab279647 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Negative control : no staining on mouse liver (PMID : 11279525).

Secondary antibody only control : Secondary antibody is ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemical analysis of paraffin-embedded mouse ovary tissue labeling Lin28A with ab279647 at 1/1000 dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining on mouse oocytes (PMID : 23378032). The section was incubated with ab279647 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse testis tissue labeling Lin28A with ab279647 at 1/100 dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution (Green). Positive staining on mouse undifferentiated spermatogonia (PMID : 32094118) is observed. The nuclear counterstain was DAPI (Blue).

Secondary antibody control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488)at 1/1000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NCCIT (human pluripotent embryonic carcinoma epithelial cell line) cells labeling Lin28A with ab279647 at 1/500 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing nucleolar and cytoplasmic staining in NCCIT cell line. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 (Red). The nuclear counterstain was DAPI (Blue).

Negative control : HEK-293T (PMID : 25479749).

Secondary antibody only control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Lin28A was immunoprecipitated from 0.35 mg F9 (mouse embryonal carcinoma epithelial cell) whole cell lysate 10 μg with ab279647 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab279647 at 1/5000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/10000 dilution.

Lane 1 : F9 whole cell lysate 10 μg

Lane 2 : ab279647 IP in F9 whole cell lysate 10 μg

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab279647 in F9 whole cell lysate

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

Exposure time : 5.5 seconds

All lanes:

Immunoprecipitation - Anti-Lin28A antibody [EPR24286-100] (ab279647)

Predicted band size: 23 kDa

Observed band size: 25 kDa

false

• IP

Supplier Data

Immunoprecipitation - Anti-Lin28A antibody [EPR24286-100] (AB279647)

Lin28A was immunoprecipitated from 0.35 mg NCCIT (human pluripotent embryonic carcinoma epithelial cell), whole cell lysate 10 μg with ab279647 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab279647 at 1/5000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/10000 dilution.

Lane 1 : NCCIT whole cell lysate 10 μg

Lane 2 : ab279647 IP in NCCIT whole cell lysate 10 μg

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab279647 in NCCIT whole cell lysate

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

Exposure time : 8 seconds

All lanes:

Immunoprecipitation - Anti-Lin28A antibody [EPR24286-100] (ab279647)

Predicted band size: 23 kDa

Observed band size: 25 kDa

false

• WB

Lab

Western blot - Anti-Lin28A antibody [EPR24286-100] (AB279647)

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

The observed MW is consistent with what has been described in the literature (PMID : 25479749)

Negative control : HEK-293T (PMID : 25479749)

Exposure time : 10 seconds

All lanes:

Western blot - Anti-Lin28A antibody [EPR24286-100] (ab279647) at 1/5000 dilution

Lane 1:

NCCIT (human pluripotent embryonic carcinoma epithelial cell) whole cell lysate at 10 µg

Lane 2:

JAR (human placenta choriocarcinoma epithelial cell) whole cell lysate at 10 µg

Lane 3:

HEK-293T (human embryonic kidney epithelial cell) whole cell lysate at 10 µg

Secondary

All lanes:

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

Predicted band size: 23 kDa

Observed band size: 25 kDa

false

• WB

Lab

Western blot - Anti-Lin28A antibody [EPR24286-100] (AB279647)

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

The observed MW is consistent with what has been described in the literature (PMID : 21962509)

Negative control : NIH/3T3 (PMID : 21962509)

Exposure time : 3 seconds

All lanes:

Western blot - Anti-Lin28A antibody [EPR24286-100] (ab279647) at 1/1000 dilution

Lane 1:

F9 (mouse embryonal carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 23 kDa

Observed band size: 25 kDa

false