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Western blot
ImmunocytochemistryAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [2]
- Other assay [1]
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- Product number
- LF-PA0048 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Anti-ROCK1 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- A suggested positive control for this product is HepG2 cells.
- Reactivity
- Human, Mouse, Rat
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µL
- Concentration
- Lot Dependent
- Storage
- -20° C, Avoid Freeze/Thaw Cycles
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of ROCK1 was achieved by transfecting PC-3 cells with ROCK1 specific validated siRNA (Silencer® select Product # s12097). Western blot analysis (Fig. a) was performed using membrane enriched extracts from the ROCK1 knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Anti-ROCK1 Polyclonal Antibody (Product # LF-PA0048, 1:1000 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). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to ROCK1.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis was performed on membrane enriched extracts (30 µg lysate) of A549 (Lane 1), PC-3 (Lane 2), Hep G2 (Lane 3) and tissue extracts of Mouse Lung (Lane 4) and Mouse Testis (Lane 5). The blot was probed with Anti-ROCK1 Polyclonal Antibody (Product # LF-PA0048, 1:1000 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 ~160 kDa band corresponding to ROCK1 was observed across the cell lines tested.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of ROCK1 was performed using 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with ROCK1 Rabbit Polyclonal Antibody(Product # LF-PA0048) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing Golgi and membrane localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
