- PA5-27421 - Invitrogen Antibodies RAB1A antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of RAB1A was performed by separating 50 µg of mouse tissue extract by 12% SDS-PAGE. Proteins were transferred to a membrane and probed with a RAB1A Polyclonal Antibody (Product # PA5-27421) at a dilution of 1:1000. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot analysis of RAB1A expression in transfected 293T cell line by RAB1A polyclonal antibody. Lane A: Non-transfected lysate. Lane B: RAB1A transfected lysate. 12% SDS PAGE. RAB1A Polyclonal Antibody (Product # PA5-27421) diluted at 1:500.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of RAB1A was achieved by transfecting U-87 MG with RAB1A specific siRNAs (Silencer® select Product # S229382, S11658). Western blot analysis (Fig. a) was performed using whole cell extracts from the RAB1A knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with RAB1A Polyclonal Antibody (Product # PA5-27421, 1:2000) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20,000). 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 RAB1A.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of RAB1A was performed by separating 50 µg of rat tissue extract by 12% SDS-PAGE. Proteins were transferred to a membrane and probed with a RAB1A Polyclonal Antibody (Product # PA5-27421) at a dilution of 1:500. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using RAB1A Polyclonal Antibody (Product # PA5-27421) and a 22 kDa band corresponding to RAB1A was observed, along with uncharacterized bands (*) around 32 kDa in Fig a and 80 & 160 kDa in Fig b. For Fig a, whole cell extracts (30 µg lysate) of HeLa (Lane 1), Caco-2 (Lane 2), BeWo (Lane 3), Hep G2 (Lane 4), U-87 MG (Lane 5), U-2 OS (Lane 6), MCF7 (Lane 7) and for Fig b, tissue extracts (30 µg lysate) of Mouse Brain (Lane 1), Rat Brain (Lane 2) were electrophoresed using NuPAGE™ 10% Bis-Tris Protein Gel (Product # NP0302BOX), 12 well. 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) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20,000) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescent detection was performed using SuperSignal™ West Pico PLUS Chemiluminescent Substrate (Product # 34580).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of RAB1A was performed by separating 30 µg of various whole cell extracts by 12% SDS-PAGE. Proteins were transferred to a membrane and probed with a RAB1A Polyclonal Antibody (Product # PA5-27421) at a dilution of 1:500 and a HRP-conjugated anti-rabbit IgG secondary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockout of RAF1 was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042, Assay ID CRISPR627776_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of RAF1 was performed by loading 30 µg of HeLa wild type (Lane 1), HeLa Cas9 (Lane 2) and HeLa RAF1 KO (Lane 3) whole cell extracts. The samples were electrophoresed using NuPAGE™ Novex™ 4-12% Bis-Tris Protein 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 c-Raf Polyclonal Antibody (Product # PA5-29333, 1:2000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20,000 dilution) using the iBright™ FL1500 (Product # A44115). Chemiluminescent detection was performed usingSuperSignal™ West Dura Extended Duration Substrate (Product # 34076). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to RAF1.

PA5-27421