- 703632 - Invitrogen Antibodies SLIRP antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of SLIRP was achieved by transfecting MCF7 cells with SLIRP specific siRNA (Silencer® select Product # s37869 & s37870). Western blot analysis (Fig a) was performed using modified whole cell extracts (1% SDS) from SLIRP knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with Anti-SLIRP Recombinant Rabbit Monoclonal Antibody (Product # 703632, 1:5000 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 knockdown confirms that antibody is specific to SLIRP.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on modified whole cell extracts (1% SDS) (30 µg lysate) of MCF7 (Lane 1), U-2 OS (Lane 2), MDA-MB-231 (Lane 3), T-47D (Lane 4), HCT 116 (Lane 5), LNCaP (Lane 6) and Hep G2 (Lane 7). The blot was probed with Anti-SLIRP Recombinant Rabbit Monoclonal Antibody (Product # 703632, 1:5000 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 ~12 kDa band corresponding to SLIRP was observed across the cell lines tested.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: For immunofluorescence analysis, MCF7 cells were fixed and permeabilized for detection of endogenous SLIRP using Anti-SLIRP Recombinant Rabbit Monoclonal Antibody (Product # 703632, 1:100 dilution) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of SLIRP protein (green), Panel b) is stained for nuclei (blue) using ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of Panels a, b and c clearly demonstrating mitochondrial localization of SLIRP. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: For immunofluorescence analysis, MCF7 cells were fixed and permeabilized for detection of endogenous SLIRP using Anti-Citrate synthase Recombinant Rabbit Monoclonal Antibody (Product # 703632, 1:100 dilution) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of SLIRP protein (green), Panel b) is stained for nuclei (blue) using ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). Panel c) represents mitochondrial staining using MitoTracker® Red CMXRos (Product # M7512). Panel d) is a composite image of Panels a, b and c clearly demonstrating co-localization of SLIRP with mitotracker which specifically binds to the mitochondria. Panel e) represents control cells without primary antibody to assess background. The images were captured at 60X magnification.

703632