Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture)

• FuncS (Neut/Block)

Supplier Data

Functional Studies (Neut/Block) - Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture) (AB244788)

Modulation of STAT1/2 phosphorylation in A431 cells following stimulation with IL-29 (ab200263), in the presence of either an anti-IL-29 antibody (ab244788) or an IgG isotype control (ab199376) using AlphaLISA SureFire assay technology.

(A) Dose–response curves of A431 cells exposed to increasing concentrations of IL-29 for 30 minutes.

(B) Analysis of phospho-STAT1 (Tyr701)/phospho-STAT2 (Tyr690) complex induced by 50ng/ml human IL-29 following treatment with anti-IL-29 antibody (ab200263; filled squares, solid line) or IgG isotype control (ab199376; open squares, dotted line) across a range of concentrations. The IC₅₀ for inhibition of phospho-STAT1 (Tyr701)/phospho-STAT2 (Tyr690) complex is 0.12 μg/mL.

(C) Analysis of phospho-STAT1 (Tyr701) induced by 50ng/ml human IL-29 following treatment with anti-IL-29 antibody (ab200263; filled triangles, solid line) or IgG isotype control (ab199376; open triangles, dotted line) across a range of concentrations. The IC₅₀ for inhibition of phospho-STAT1 (Tyr701) is 0.11 μg/mL.

(D) Analysis of phospho-STAT2 (Tyr690) induced by 50ng/ml human IL-29 following treatment with anti-IL-29 antibody (ab200263; filled circles, solid line) or IgG isotype control (ab199376; open circles, dotted line) across a range of concentrations. The IC₅₀ for inhibition of phospho-STAT2 (Tyr690) is 0.13 μg/mL.

A431 cells were seeded at 4 × 10⁴ cells per well in 96‑well plates. Anti‑IL‑29 antibody or isotype control at the indicated concentrations was pre‑incubated with 50 ng/mL IL‑29 for 10 minutes before being added to the cells, followed by a 30‑minute stimulation. Levels of phospho‑STAT1 (Tyr701), phospho‑STAT2 (Tyr690), and the phospho‑STAT1 (Tyr701)/phospho‑STAT2 (Tyr690) complex were quantified using SureFire^® Ultra™ assays.

The data were kindly generated by the TGR Biosciences team, an abcam company SureFire^® is a registered trademark of TGR BioSciences Pty Ltd, Australia.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture) (AB244788)

Flow cytometry (intracellular) analysis of transfected 293T cells. Cells were fixed with 4% PFA and permeabilised with 90% methanol before staining for IL-29 in 293T cells transfected with a human IFNL1 expression vector containing a myc-His-tag^® (Middle) using ab244788 at 1/50000 dilution (0.001ug). Staining was compared with 293T cells transfected with an empty expression vector containing a myc-His-tag^® (Right) and with an isotype control, Rabbit monoclonal IgG (ab172730) (Left). Goat anti‑rabbit IgG (Alexa Fluor^® 488, ab150081) was used as the secondary antibody at a 1/5000 dilution. Cells were co-stained with Myc tag conjugated to Alexa Fluor^® 647.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture) (AB244788)

Immunofluorescent analysis of 4% Paraformaldehyde fixed 0.1% Triton X-100 permeabilized HEK-293T cells transfected with a human IL-29/IFNL1 protein expression vector containing a myc-His-tag^® labelling IL-29 with ab244788 at 1/1000 (1.043 μg/ml) dilution. ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed was used as secondary at 1/1000 (2μg/ml) dilution. ab223894 Anti-Myc tag mouse monoclonal antibody (Alexa Fluor^® 594) was used as a counterstain at 1/200 dilution (2.5μg/ml). Confocal image showing positive staining in 293T cells (shown in green) transfected with a human IFNL1 protein expression vector containing a myc-His-tag^®. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).

Negative control : HEK-293T cells (human embryonic kidney epithelial cells) transfected with an empty expression vector containing a myc-His-tag^®.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture) (AB244788)

Immunohistochemical analysis of HEK-293T (human embryonic kidney epithelial cell) transfected with a human IFNL1 expression vector containing a myc-His-tag labeling IL-29 with ab244788 at 1/5000 (0.209 μg/ml) dilution. Positive staining on (A) HEK-293T transfected with a human IFNL1 expression vector containing a myc-His-tag, no staining on (B) HEK-293T transfected with empty vector containing a myc-His-tag. The primary antibody was incubated for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• sELISA

Supplier Data

Sandwich ELISA - Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture) (AB244788)

Representative standard curve from corresponding SimpleStep ELISA® Kit (ab236715).

• WB

Lab

Western blot - Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture) (AB244788)

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

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, 6X His tag® antibody [AD1.1.10]- Total Protein Control (ab15149) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-IL-29 antibody [EPR19773-96] - BSA and Azide free (Capture) (ab244788) at 1/1000 dilution

Lane 1:

293T cells transfected with an empty vector containing a His-tag, whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human IFNL1 expression vector containing a His-tag, 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

Observed band size: 25 kDa,36 kDa

false

Exposure time: 15s