Anti-FOXP1 antibody [SP133] - C-terminal

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Flow cytometry staining of human peripheral blood mononuclear cells (PBMCs) with ab227649 (right) or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (left). Cells were fixed and permeabilised with BD Cytofix/Cytoperm™ for 20 min. PBMCs were incubated for 30 min at 4°C in 1x PBS containing 10 μg/ml human IgG and 10 % normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody ab227649 or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (1x 106 in 100 μl at 0.2 μg/ml (1/10050)) for 30 min at 4°C . The cells were simultaneously stained with CD4.

The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 4°C

Acquisition of >30000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter. Events were gated on viable cells.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Flow cytometric analysis of MOLT-4 (human lymphoblastic leukemia cell line) cell line labeling FOXP1 with ab227649 at 1/100 dilution (green) compared with a negative control of rabbit IgG (blue).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human tonsil tissue sections labeling FOXP1 with ab227649 at 1/100 dilution (2.40 µg/ml). Heat mediated antigen retrieval was performed Heat mediated antigen retrieval with sodium citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Formalin-fixed, paraffin-embedded human tonsil tissue stained for FOXP1 using ab227649 at 1/100 dilution in immunohistochemical analysis.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

ab227649 staining FOXP1 in A549 (positive) and A549-FOXP1 (negative) cells. The cells were fixed with 100% methanol (5 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 for 2 hours at room temperature with ab227649 at 1ug/ml (shown in green) and ab7291, Anti-alpha Tubulin antibody [DM1A] - Loading Control (shown in pseudocolour magenta). Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 dilution and ab150119, Goat Anti-Mouse IgG H&L (Alexa Fluor^® 647) preadsorbed at 1/1000 dilution for 1 hour at room temperature. Nuclear DNA was labeled with DAPI (shown in blue). Also suitable in cells fixed with 4% paraformaldehyde (10 min). Image was acquired with a high-content analyser (Operetta CLS, Revvity) and a maximum intensity projection of confocal sections is shown.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Flow cytometry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) labeling FOXP1 with purified ab227649 at 1/20 dilution (12 μg/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as a secondary antibody. Isotype control -Rabbit monoclonal IgG (ab172730) / Black. Unlableled control -Unlabelled cells / blue.

• IHC-Fr

Unknown

Immunohistochemistry (Frozen sections) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Immunohistochemistry (Frozen) analysis of rat cerebral cortex tissue section labeling FOXP1 with purified ab227649 at 1/25 (9.6 μg/ml). Sections were fixed in 4% paraformaldehyde and permeabilized with 0.2% Triton X-100. Antigen retrieval was Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 μg/ml) dilution. DAPI was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Rat colon tissue sections labeling FOXP1 with ab227649 at 1/400 dilution (0.60 µg/ml). Heat mediated antigen retrieval was performed Heat mediated antigen retrieval with sodium citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• IHC-Fr

Unknown

Immunohistochemistry (Frozen sections) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Immunohistochemistry (Frozen) analysis of rat cerebrum tissue section labeling FOXP1 with purified ab227649 at 1/25 (9.6 μg/ml). Sections were fixed in 4% paraformaldehyde and permeabilized with 0.2% Triton X-100. Antigen retrieval was Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20). Goat anti rabbit IgG (Alexa Fluor ^®488, ab150077) was used as the secondary antibody at 1/1000 (2 μg/ml) dilution. DAPI was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Mouse colon tissue sections labeling FOXP1 with ab227649 at 1/400 dilution (0.60 µg/ml). Heat mediated antigen retrieval was performed Heat mediated antigen retrieval with sodium citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• WB

Lab

Western blot - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

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

Lysate was freshly made and used for Western blotting immediately to minimize protein degradation.

ab227649 is less sensitive than ab314488 in WB. We recommend using ab314488 for WB testing.

All lanes:

Western blot - Anti-FOXP1 antibody [EPR26484-79] (<a href='/en-us/products/primary-antibodies/foxp1-antibody-epr26484-79-ab314488'>ab314488</a>) at 1/1000 dilution

All lanes:

HEK 293 (human embryonic kidney epithelial cell) whole cell lysate at 20 µg with NFDM/TBST

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/20000 dilution

Observed band size: 90 kDa,70 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-FOXP1 antibody [SP133] - C-terminal (AB227649)

All lanes:

Western blot - Anti-FOXP1 antibody [SP133] - C-terminal (ab227649) at 1/25 dilution

All lanes:

MCF7 (human breast adenocarcinoma cell line) cell lysate

Predicted band size: 75 kDa

false