- MA5-32111 - Invitrogen Antibodies BAK1 antibody

Submitted references Inflammation-dependent downregulation of miR-532-3p mediates apoptotic signaling in human sarcopenia through targeting BAK1.

Chen FX, Shen Y, Liu Y, Wang HF, Liang CY, Luo M
International journal of biological sciences 2020;16(9):1481-1494

Intramuscular accumulation of pentadecanoic acid activates AKT1 to phosphorylate NCOR1 and triggers FOXM1-mediated apoptosis in the pathogenesis of sarcopenia.

Chen FX, Du N, Hu J, Ning F, Mei X, Li Q, Peng L
American journal of translational research 2020;12(9):5064-5079

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of BAK in different lysates using a Monoclonal antibody (Product #MA5-32111) at a dilution of 1:1,000. Positive control: Lane 1: Hela, Lane 2: Human skeletal muscle, Lane 3: Ags.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of BAK in different lysates using a Monoclonal antibody (Product #MA5-32111) at a dilution of 1:1,000. Positive control: Lane 1: Hela, Lane 2: Human skeletal muscle, Lane 3: Ags.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-BAK Recombinant Rabbit Monoclonal Antibody (SU32-07) (Product # MA5-32111) and a 25 kDa band corresponding to BAK1 was observed across cell lines tested. Membrane enriched extracts (30 µg lysate) of MCF7 (Lane 1), Ramos (Lane 2), K-562 (Lane 3) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) 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 # A27036,1:5000 dilution) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescentdetection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of BAK1 was achieved by transfecting MCF7 with BAK1 specific siRNAs (Silencer® select Product # S1880, S1879). Western blot analysis (Fig. a) was performed using Membrane enriched extracts from the BAK1 untransfected cells (lane 1), non-targeting scrambled siRNA transfected cells (lane 2) and knockdown cells (lane 3). The blot was probed with BAK Recombinant Rabbit Monoclonal Antibody (SU32-07) (Product # MA5-32111, 1:1000 dilution ) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:5000 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 BAK1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunocytochemical analysis of BAK in CRC cells using a BAK Monoclonal antibody (Product # MA5-32111) as seen in green. The nuclear counter stain is DAPI (blue). Cells were fixed in paraformaldehyde, permeabilised with 0.25% Triton X100/PBS.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of BAK1 was performed using 70% confluent log phase A-431 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.01% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with BAK Recombinant Rabbit Monoclonal Antibody (SU32-07) (Product # MA5-32111) 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). 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 mitochondria and cell membrane localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 40X magnification in CellInsight CX7 LZR High-Content Screening (HCS) Platform (Product # CX7A1110LZR) and externally deconvoluted (D.Sage et al. / Methods 115 (2017) 28–41).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of BAK of paraffin-embedded Human lung tissue using a BAK Monoclonal antibody (Product #MA5-32111). Counter stained with hematoxylin.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical analysis of BAK of paraffin-embedded Mouse lung tissue using a BAK Monoclonal antibody (Product #MA5-32111). Counter stained with hematoxylin.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow Cytometric analysis of BAK in NIH/3T3 cells using a BAK Monoclonal Antibody (Product # MA5-32111) at a dilution of 1:50, as seen in blue compared with an unlabelled control (cells without incubation with primary antibody; red). Alexa Fluor 488-conjugated goat anti rabbit IgG was used as the secondary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 LPS treatment could affect the expression of miR-532-3p and BAK1 and their downstream molecules. (A) Serum levels of BAK1 were significantly elevated. Serum samples from 24 healthy and 24 sarcopenia patients were used to measure circulating BAK1 concentrations using ELISA. *** P < 0.001. (B) BAK1 and its downstream caspase cascades were activated in sarcopenia samples. Three-paired muscle tissues from healthy controls and sarcopenia patients were used to detect the protein levels of BAK1, Caspase-3 and Caspase-9. GAPDH was used as a loading control. F: full length; C: cleaved length. (C) LPS induced the expression of BAK1 . Three cell lines, HSMM-1, HSMM-2 and LHCN-M2, were treated with 200 ng/mL LPS for 6 h, followed by RNA isolation and qRT-PCR analyses to measure the mRNA level of BAK1 . *** P < 0.001. (D) LPS induced the BAK1 protein level and its downstream caspase cascades. The cells used in (C) were subjected to protein extraction, followed by the detection of the BAK1, Caspase-3 and Caspase-9 protein levels. GAPDH was used as a loading control. (E-G) The combined effects of miR-532-3p-mimic/anti-miR-532-3p and LPS treatment. The HSMM-1 and HSMM-2 cells were primarily transfected with miR-532-3p-mimic or anti-miR-532-3p. After 48 h, cells were treated with 200 ng/mL LPS for 6 h and then were collected and subjected to RNA isolation and protein extraction. The resulting RNA and protein extracts used for the detection of the expression of miR-532-3p (E) , BAK1 m

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