- PA5-29039 - Invitrogen Antibodies MCM4 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of MCM4 using 30 µg of MOLT4 lysate. Samples were loaded onto a 7.5% SDS-PAGE gel and probed with a MCM4 polyclonal antibody (Product # PA5-29039) at a dilution of 1:5000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot analysis of MCM4 was performed by separating 30 µg of various whole cell extracts by 7.5% SDS-PAGE. Proteins were transferred to a membrane and probed with a MCM4 Polyclonal Antibody (Product # PA5-29039) at a dilution of 1:1,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: MCM4 Polyclonal Antibody detects MCM4 protein by western blot analysis. A. 30 µg Jurkat whole cell lysate/extract. B. 30 µg Raji whole cell lysate/extract. C. 30 µg NCI-H929 whole cell lysate/extract.7.5 % SDS-PAGE. MCM4 Polyclonal Antibody (Product # PA5-29039) dilution: 1:5,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of MCM4 was performed by separating 30 µg of various whole cell extracts by 7.5% SDS-PAGE. Proteins were transferred to a membrane and probed with a MCM4 Polyclonal Antibody (Product # PA5-29039) at a dilution of 1:1,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: MCM4 Polyclonal Antibody detects MCM4 protein by western blot analysis. A. 30 µg Jurkat whole cell lysate/extract. B. 30 µg Raji whole cell lysate/extract. C. 30 µg NCI-H929 whole cell lysate/extract.7.5 % SDS-PAGE. MCM4 Polyclonal Antibody (Product # PA5-29039) dilution: 1:5,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of MCM4 was achieved by transfecting SH-SY5Y with MCM4 specific siRNAs (Silencer® select Product #sS8592, s8593). Western blot analysis (Fig. a) was performed using nuclear enriched extracts from the MCM4 knockdown cells (Lane 1), non-specific scrambled siRNA transfected cells (Lane 2), untransfected cells (Lane 3), non-targeted siRNA (Lane 4). The blot was probed with MCM4 Polyclonal Antibody (Product # PA5-29039, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 0.25µg/ml, 1:4000 dilution). Densitometric analysis of this western blot is shown i histogram (Fig. b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to MCM4.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using MCM4 Polyclonal Antibody (Product # PA5-29039) and a 110 kDa band corresponding to MCM4 was observed across cell lines tested. Whole cell extracts (30 µg lysate) of Neuro-2a (Lane 1), NIH/3T3 (Lane 2), NIH: OVCAR-3 (Lane 3), U-2 OS (Lane 4), SH-SY5Y (Lane 5) were electrophoresed using NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX). 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, 1:4000 dilution) (Product # A27014) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of MCM4 in paraformaldehyde-fixed HeLa cells using a MCM4 polyclonal antibody (Product # PA5-29039) at a 1:500 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of MCM4 was performed using 70% confluent log phase SH-SY5Y 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 1 hour at room temperature. The cells were labeled with MCM4 Polyclonal Antibody (Product # PA5-29039) at 5 µg/mL 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) 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 nuclear localization. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of paraffin-embedded A549 Xenograft, using MCM4 (Product # PA5-29039) antibody at 1:100 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Cross-linked ChIP was performed with HeLa chromatin extract and 5 µg of either control rabbit IgG or a MCM4 polyclonal antibody (Product # PA5-29039). The precipitated DNA was detected by PCR with primer set targeting to Myc promoter.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Cross-linked ChIP was performed with HeLa chromatin extract and 5 µg of either control rabbit IgG or MCM4 Polyclonal Antibody (Product # PA5-29039). The precipitated DNA was detected by PCR with primer set targeting to Myc promoter.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Chromatin Immunoprecipitation (ChIP) assay of endogenous MCM4 protein using Anti-MCM4 Antibody: ChIP was performed using Anti-MCM4 Rabbit Polyclonal Antibody (Product # PA5-29039) 5 µg, on sheared chromatin from HeLa cells using the MAGnify ChIP System kit (Product # 49-2024). Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR using primers binding to transcriptional start site and gene body (+2kb) of GAPDH, transcriptional start site of MYC and SAT2 satellite repeats. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: MCM4 Polyclonal Antibody immunoprecipitates MCM4 protein in IP experiments. IP samples: Jurkat whole cell extract. A. 40 µg Jurkat whole cell extract. B. Control with 4 µg of preimmune Rabbit IgG. C. Immunoprecipitation of MCM4 protein by 4 µg MCM4 Polyclonal Antibody (Product # PA5-29039). 7.5 % SDS-PAGE. The immunoprecipitated MCM4 protein was detected by MCM4 Polyclonal Antibody (Product # PA5-29039) diluted at 1:500.

PA5-29039