- MA5-14991 - Invitrogen Antibodies ERN1 antibody

Guo Y, Hao D, Hu H
Molecular biology reports 2021 Dec;48(12):7841-7851

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis was performed on whole cell extracts (30 µg lysate) of Hep G2 (Lane 1), PANC-1 (Lane 2), Raji (Lane 3), Neuro-2a (Lane 4) and HEK293T (Lane 5). The blot was probed with Anti-IRE1 alpha Monoclonal Antibody (J.607.10) (Product # MA5-14991, 1:500 dilution) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). A 120 kDa band corresponding to IRE1 alpha was observed in all the cell lines tested.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of IRE1 alpha was achieved by transfecting PANC-1 with IRE1 alpha specific siRNAs (Silencer® select Product # s200430). Western blot analysis (Fig. a) was performed using whole cell extracts from the IRE1 alpha knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with IRE1 alpha Monoclonal Antibody (J.607.10) (Product # MA5-14991, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to IRE1 alpha.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: RNA immunoprecipitation (RIP) western of ERN1 was performed in K562 cells. Antigen-antibody complexes were formed by incubating approximately 500 µg whole cell lysate with 5 to 10 µL of monoclonal ERN1 antibody (Product # MA5-14991) rotating 60 min at RT. The immune complexes were captured on 625 µg of anti-rabbit coated Dynabeads (Product # 11204D) and washed extensively. They were then eluted and analyzed using the Simple Western system using the same antibody as used in immunoprecipitation at a dilution of 1:25, followed by a 1:100 dilution of secondary antibody. Lane 1 is the input, lane 2 no antibody IP and lane 3 is the target specific IP. Data courtesy of the Yeo lab as part of the ENCODE project.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: RNA immunoprecipitation (RIP) western of ERN1 was performed in K562 cells. Antigen-antibody complexes were formed by incubating approximately 500 µg whole cell lysate with 5 to 10 µL of monoclonal ERN1 antibody (Product # MA5-14991) rotating 60 min at RT. The immune complexes were captured on 625 µg of anti-rabbit coated Dynabeads (Product # 11204D) and washed extensively. They were then eluted and analyzed using the Simple Western system using the same antibody as used in immunoprecipitation at a dilution of 1:25, followed by a 1:100 dilution of secondary antibody. Lane 1 is the input, lane 2 no antibody IP and lane 3 is the target specific IP. Data courtesy of the Yeo lab as part of the ENCODE project.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 1 MC3T3-E1 cells were treated with Dex (0, 10 -8 , 10 -6 , and 10 -4 M) for 24 h. a Western blot of ATF6, phosphorylated PERK (p-PERK), PERK, phosphorylated IRE1 (p-IRE1), and IRE1 in MC3T3-E1 cells. b TUNEL staining was used to detect the effect of different concentrations of Dex on apoptosis in MC3T3-E1 cells (Scale=50 mum); c Caspase-12 activity was measured in cells receiving Dex; d Caspase-3 activity was measured in cells receiving Dex

MA5-14991