Anti-FOXP3 antibody [236A/E7] - BSA and Azide free

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

This data was developed using the same antibody clone in a different buffer formulation (ab20034).

Immunohistochemical analysis of formalin fixed paraffin embedded human tonsil labelling FOXP3 with ab20034 at a concentration of 2 µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with an OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was conducted for 32 mins at 100°C with ULTRA cell conditioning solution (CC1, pH 8.5). ab20034 anti-FOXP3 antibody [236A/E7] was incubated at 37°C for 16 mins. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data is courtesy of ImmunoAtlas and it can be found here.

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

This image is courtesy of ImmunoAtlas.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

This data is courtesy of ImmunoAtlas and it can be found here.

This image is courtesy of ImmunoAtlas.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data is courtesy of ImmunoAtlas and it can be found here.

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

This image is courtesy of ImmunoAtlas.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

This data is courtesy of ImmunoAtlas and it can be found here.

This image is courtesy of ImmunoAtlas.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

This data was developed using the same antibody clone in a different buffer formulation (ab20034).

Immunohistochemical analysis of formalin fixed paraffin embedded) human tonsil labelling FOXP3 with ab20034 at a concentration of 2 µg/ml. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with a Bond™ Polymer Refine Detection kit. Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution 2) for 20 mins. ab20034 anti-FOXP3 antibody [236A/E7] was incubated for 30 mins at room temperature. Sections were counterstained with Hematoxylin. Image inset shows absence of staining in secondary antibody only control.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

This data is courtesy of ImmunoAtlas and it can be found here.

This image is courtesy of ImmunoAtlas.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Human tonsil tissue labeling FOXP3 with ab23004 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Counterstained with hematoxylin. Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat AntiRabbit IgG H&L (HRP). Perform heat mediated antigen retrieval using Tris/EDTA buffer (pH 9.0).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Human thymus tissue labeling FOXP3 with ab23004 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Counterstained with hematoxylin. Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat AntiRabbit IgG H&L (HRP). Perform heat mediated antigen retrieval using Tris/EDTA buffer (pH 9.0).

• WB

Supplier Data

Western blot - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

All lanes:

Western blot - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (ab96048) at 1/1000 dilution

Lane 1:

HEK-293 (human embryonic kidney) transfected with an empty vector (vector control), containing a GFP-Myc-tag, whole cell lysate at 10 µg

Lane 2:

HEK-293 transfected with FOXP3 (WT) expression vector containing a GFP-Myc-tag, whole cell lysate at 10 µg

Predicted band size: 47 kDa

false

Exposure time: 1s

• WB

Supplier Data

Western blot - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

All lanes:

Western blot - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (ab96048) at 1/1000 dilution

All lanes:

Human mammary gland lysate at 20 µg

Predicted band size: 47 kDa

false

• WB

Supplier Data

Western blot - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (AB96048)

This data was developed using ab20034, the same antibody clone in a different buffer formulation.

ab20034 detects Human FOXP3 protein at ~50 kDa in HEK293T cells overexpressing the protein. It also detects FOXP3 in Human tonsil tissue lysate, however this band is significantly weaker in endogenous conditions. Upon higher exposure, weak bands can also be observed in HEK293T cell lysate.

This blot was produced using a 4-12% Bis-Tris gel under the MOPS buffer system. The gel was run at 200V for 60 minutes before being transferred onto a nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour before being incubated with Anti-FOXP3 antibody [236A/E7] (ab20034; 5 microgram per mL) overnight at 4°C. Antibody binding was detected using infrared labelled goat anti-mouse (green; 1 : 10000) for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-FOXP3 antibody [236A/E7] - BSA and Azide free (ab96048) at 5 µg/mL

Lane 1:

HEK293T cell lysate at 20 µg

Lane 2:

HEk293T cell lysate overexpressing Human FOXP3 at 20 µg

Lane 3:

Human tonsil tissue lysate at 20 µg

Predicted band size: 47 kDa

false