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CDK5 Antibody Cocktail
Antibody target was verified by KnockDown to ensure the antibody binds to the antigen stated.
Catalog # MA5-11291
(USD) 349.00, 500 µL
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Immunofluorescence analysis of Cdk5 was done on 70% confluent log phase MCF-7 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Cdk5 (DC17 + DC34) Mouse Monoclonal Antibody (Product # MA5-11291) at 2 µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) 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 Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d is a merged image showing cytoplasmic localization. Panel e is a no primary antibody control. The images were captured at 60X magnification.
Western blot analysis of CDK5 was performed with 10 µg of HeLa cells transfected with Transfection Reagent alone (Lane 1), 100nM Non-Targeting control siRNA (Lane 2), or 100nM siRNA against CDK5 (Lane 3). Proteins were resolved using a NuPAGE® Novex 4-12% Bis-Tris Gel (Product # NP0322BOX), XCell SureLock™ Electrophoresis System (Product # EI0002), and a protein size ladder. Proteins were wet transferred to a Pierce Nitrocellulose Membrane (Product # 88025) OR Pierce PVDF Membrane (Product # 88518) and blocked with Pierce Starting Block T20 (PBS) Blocking Buffer (Product # 37539) for 1 hour at room temperature. CDK5 was detected at ~ 31 kDa using CDK5 Mouse monoclonal antibody (Product # MA5-11291) diluted in Pierce Starting Block T20 (PBS) Blocking Buffer 4°C overnight on a rocking platform. Pierce Goat Anti-Mouse (Product # 31437) HRP-Conjugated Antibodies at a 1:2500 dilution were used and chemiluminescent detection was performed using Pierce Supersignal West Dura Maximum Sensitivity Substrate (Product # 37071). Relative density of the bands normalized to GAPDH (36 kDa). CDK5 Antibody (Product # MA5-11291) confirms silencing of CDK5 expression.
Western blot analysis was performed on whole cell extracts (30 µg lysate) of PC-3 (lane 1), SH-SY5Y (lane 2), PC-12 (lane 3), Jurkat (lane 4) and U-87 MG (lane 5). The blots were probed with Anti-Cdk5 Mouse Monoclonal Antibody (Product # MA5-11291, 1-2 µg/mL) and detected by chemiluminescence oat anti-Mouse IgG (H+L) Secondary Antibody, HRP conjugate (Product # 62-6520, 1:4000 dilution). A 33 kDa band corresponding to Cdk5 was observed across cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody using iBind™ Flex Western Starter Kit (Product # SLF2000S). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
Western blot of Cdk5 using Cdk5 Monoclonal Antibody (Product # MA5-11291) on IMR-5 Cells.
Immunoprecipitation and Western blot of Cdk5 was performed using Jurkat cell lysates. Cells were lysed using ice cold IP lysis/wash buffer and pre-cleared using 50 ml of bead slurry per ml of cell lysate. Antigen-antibody complexes were formed by incubating 0.5 ml pre-cleared cell lysate on ice for 3hrs with 8-15ug of Cdk5 monoclonal antibody (Product # MA5-11291) crosslinked to Protein A/G plus agarose. The immune complexes were eluted using 60 ul sample buffer boiled at 95°C for 5 min and loaded onto an SDS-PAGE gel; Input (lane 1), Jurkat IP (Lane 2) and Jurkat supernatant (lane 3). The membrane was probed with a Cdk5 monoclonal antibody (Product # MA5-11291) at a dilution of 6ug/mg of lysate followed by detection using an HRP-conjugated goat anti-mouse IgG + IgM (H+L) cross-adsorbed secondary antibody. Chemiluminescent detection was performed using an exposure time of 20s, resulting in a ~30kDa band on input and IP lysates.
Immunoprecipitation of Cdk5 using Cdk5 Monoclonal Antibody (Product # MA5-11291) on Native Human LS174T Cells.
Antibody specificity was demonstrated by siRNA mediated knockdown of target protein. HeLa cells were transfected with CDK5 siRNA and decrease in signal intensity was observed in western blot application using anti-CDK5 monoclonal Antibody (Product # MA5-11291). Densitometric analysis of this western blot is shown in histogram (Fig b).
6 ug/mg protein lysate
Western Blot (WB)
Host / Isotype
DC17 + DC34
Recombinant cdk5 protein
PBS, pH 7.4, with 0.2% BSA
0.09% sodium azide
Product Specific Information
MA5-11291 targets Cdk5 in IF, IP, and WB applications and shows reactivity with Human, mouse, and Rat samples.
The MA5-11291 immunogen is recombinant cdk5 protein.
Cdk5 is expressed in both cycling and noncycling cells and is present in numerous mammalian tissues. Neuronal and muscle cells contain the highest amount of this protein. The p35 protein, which is expressed solely in the brain, activates Cdk5. Cdk5 activity is involved in terminal differentiation of neurons and muscle cells.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
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