Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [1]
- Immunocytochemistry [2]
- Immunohistochemistry [1]
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Validation data
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- Product number
- PA5-38297 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Anti-Phospho-MYPT1 (Thr696) Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Reactivity
- Human, Mouse, Rat
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µg
- Concentration
- 1 mg/mL
- Storage
- -20°C
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-Phospho-MYPT1 (Thr696) Polyclonal Antibody(Product # PA5-38297) and a 110kDa band corresponding to Phospho-MYPT1 (Thr696) was observed across the cell lines tested. Whole cell extracts (30 µg lysate) of HEK 293 (Lane 1) and NTERA-2 cl.D1 (Lane 2), which were lambda phosphatase treated and HEK 293 (Lane 3) and NTERA-2 cl.D1 (Lane 4), which were not lambda phosphatase treated, were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # LC2001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036,1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).In presence of alkaline phosphatase treatment, there were no bands observed but in the untreated lanes the bands can be observed, which shows that the SKU is specific to Phosphorylation sites.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent analysis of Phospho-MYPT1 pThr696 in A549 cells using a Phospho-MYPT1 pThr696 polyclonal antibody (Product # PA5-38297).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Phospho-MYPT1 (Thr696) was performed using 70% confluent log phase HEK-293 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with Phospho-MYPT1 (Thr696) Polyclonal Antibody (Product # PA5-38297) 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™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034), (1:2000 dilution), for 45 minutes at room temperature (Panel a: Green, treated with 10 uM Calyculin A for 10 mins). Nuclei (Panel b:Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300 dilution). Panel d represents the merged image showing Cytoskeleton and cytosolic localization. Panel e represents untreated cells with low expression of the protein. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X with Oil immersion magnification.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemical analysis of Phospho-MYPT1 pThr696 in paraffin-embedded human breast carcinoma using a Phospho-MYPT1 pThr696 polyclonal antibody (Product # PA5-38297).