重组Anti-Indoleamine 2，3-dioxygenase抗体[SP260] - C-terminal (ab228468)

Western blot: Anti-IDO1 antibody [SP260] (ab228468) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab228468 was shown to bind specifically to IDO1. A band was observed at 42 kDa in treated wild-type A549 cell lysates with no signal observed at this size in IDO1 knockout cell line. To generate this image, wild-type and IDO1 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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 IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

Immunocytochemistry/ Immunofluorescence analysis of HeLa (human cervix adenocarcinoma epithelial cell) treated with 50 ng/ml IFN-gamma for 16 h cells labeling Indoleamine 2, 3-dioxygenase with purified ab228468 at 1/15 (9.4µg/ml). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 µg/ml). 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.

Flow cytometry analysis of HeLa cells treated with 50ng/ml IFN-gamma for 16hours labeling Indoleamine 2, 3-dioxygenase with purified ab228468 at 1/200 dilution (0.705 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.Untreated cells (Green).

Formalin-fixed, paraffin-embedded human tonsil tissue stained for Indoleamine 2, 3-dioxygenase with ab228468 at 1/100 dilution in immunohistochemical analysis.

Flow Cytometry analysis of IFN-gamma treated HeLa (human epithelial cell line from cervix adenocarcinoma) cells, labeling Indoleamine 2, 3-dioxygenase with ab228468 1/400 dilution (green) compared to a negative control rabbit IgG (blue).

Formalin-fixed, paraffin-embedded human thymus tissue stained for Indoleamine 2, 3-dioxygenase with ab228468 at 1/100 dilution in immunohistochemical analysis.

Formalin-fixed, paraffin-embedded human colon tissue stained for Indoleamine 2, 3-dioxygenase with ab228468 at 1/100 dilution in immunohistochemical analysis.

Formalin-fixed, paraffin-embedded human stomach adenocarcinoma tissue stained for Indoleamine 2, 3-dioxygenase with ab228468 at 1/100 dilution in immunohistochemical analysis.

Formalin-fixed, paraffin-embedded human pancreatic adenocarcinoma tissue stained for Indoleamine 2, 3-dioxygenase with ab228468 at 1/100 dilution in immunohistochemical analysis.

Formalin-fixed, paraffin-embedded human endometrial adenocarcinoma tissue stained for Indoleamine 2, 3-dioxygenase with ab228468 at 1/100 dilution in immunohistochemical analysis.

Formalin-fixed, paraffin-embedded human ovarian adenocarcinoma tissue stained for Indoleamine 2, 3-dioxygenase with ab228468 at 1/100 dilution in immunohistochemical analysis.