Anti-OAS1 antibody [EPR24824-162]

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-OAS1 antibody [EPR24824-162] (AB272492)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized 293T (Human embryonic kidney epithelial cell, left) / Daudi (Human Burkitt's lymphoma lymphoblast, right) cells labelling OAS1 with ab272492 at 1/50 dilution (1ug)/ red (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody. Negative control : 293T(PMID : 19923450).

• IP

Lab

Immunoprecipitation - Anti-OAS1 antibody [EPR24824-162] (AB272492)

OAS1 was immunoprecipitated from 0.35 mg NCI-H460 (human Large Cell Lung Cancer epithelial cell) whole cell lysate 10 ug with ab272492 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab272492 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : NCI-H460 (human Large Cell Lung Cancer epithelial cell) whole cell lysate 10 ug

Lane 2 : ab272492 IP in NCI-H460 whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab272492 in NCI-H460 whole cell lysate

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

Exposure time : 32 seconds

All lanes:

Immunoprecipitation - Anti-OAS1 antibody [EPR24824-162] (ab272492)

Predicted band size: 46 kDa

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• WB

Lab

Western blot - Anti-OAS1 antibody [EPR24824-162] (AB272492)

Blocking buffer and concentration : Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS Diluting buffer and concentration : Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBST Lysates/proteins at 20 µg per lane. Performed under reducing conditions. False colour image of Western blot : Anti-OAS1antibody [EPR24824-162] (ab272492) staining at 1/1000 dilution, shown in green; Mouse anti-alpha Tubulin antibody [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab272492 was shown to bind specifically to OAS1. A band was observed at 42 kDa in wild-type HeLa cell lysates with no signal observed at this size in OAS1 knockout cell lysates (ab259009) (knockout cell line ab265578). To generate this image, wild-type and OAS1 knockout HeLa cell lysates were analyzed. First, samples were run on an SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS 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 (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/10000 dilution.

Lanes 1 - 4:

Western blot - Anti-OAS1 antibody [EPR24824-162] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/oas1-antibody-epr24824-162-bsa-and-azide-free-ab284864'>ab284864</a>) at 1/1000 dilution

Lanes 1 - 4:

Western blot - Anti-OAS1 antibody [EPR24824-162] (ab272492) at 1/1000 dilution

Lane 1:

Wild-type HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

Wild-type HeLa treated with 1000 U/ml IFN beta for 28 hours, whole cell lysate at 20 µg

Lane 3:

OAS1 knockout HeLa whole cell lysate at 20 µg

Lane 4:

OAS1 knockout HeLa treated with 1000 U/ml IFN beta for 28 hours, whole cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/10000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 46 kDa

Observed band size: 42 kDa

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• WB

Lab

Western blot - Anti-OAS1 antibody [EPR24824-162] (AB272492)

Blocking and diluting buffer and concentration : 5% NFDM/TBSTThe molecular weight observed is consistent with what has been described in the literature (PMID : 25205466).

Negative control : LNCaP, 293T (PMID : 26997919).

Exposure time : Lane 1-2 : 3 minutes Lane 3-6 : 37 seconds

All lanes:

Western blot - Anti-OAS1 antibody [EPR24824-162] (ab272492) at 1/1000 dilution

Lane 1:

Untreated HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate

Lane 2:

HeLa treated with /ml IFN alpha 1 for 16 hours, whole cell lysate 20

Lane 3:

NCI-H460 (human Large Cell Lu Cancer epithelial cell) whole cell lysate 20

Lane 4:

Daudi (human Burkitts lymphoma lymphoblast) whole cell lysate

Lane 5:

LNCaP (human prostate carcinoma epithelial cell) whole cell lysate

Lane 6:

293T (human embryonic kidney epithelial cell) whole cell lysate

Secondary

All lanes:

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

Predicted band size: 46 kDa

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