- MA5-16237 - Invitrogen Antibodies TLR5 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of TLR5 in A) Ramos and B) Raw lysate using a TLR5 monoclonal antibody (Product # MA5-16237) at 2 µg/mL.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-TLR5 Monoclonal Antibody (19D759.2) (Product # MA5-16237) and a 93 kDa band corresponding to Toll-like receptor 5 was observed across the cell lines tested. Whole cell extracts (30 µg lysate) of HT-29 (Lane 1), U-937 (Lane 2), A549 (Lane 3) and NIH:OVCAR-3 (Lane 4) were electrophoresed using NuPAGE™ 10% Bis-Tris Protein Gel (Product # NP0302BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23002) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (at a concentration of 1 µg/mL) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177,1:4000 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 Toll-like receptor 5 was achieved by transfecting HT-29 with Toll-like receptor 5 specific siRNAs (Silencer® select Product # s14199, s14197). Western blot analysis (Fig. a) was performed using Whole cell extracts from the Toll-like receptor 5 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with TLR5 Monoclonal Antibody (19D759.2) (Product # MA5-16237, at a concentration of 1 µg/mL ) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 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 Toll-like receptor.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of TLR5 in Human skin. Samples were incubated in TLR5 monoclonal antibody (Product # MA5-16237) using a dilution of 5 µg/mL followed by a peroxidase-conjugate and DAB chromogen.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of TLR5 in formalin-fixed paraffin-embedded tissue section of normal human skin. Samples were incubated in TLR5 monoclonal antibody (Product # MA5-16237) using a dilution of 5 µg/mL. Intense membrane staining of TLR5 was observed specifically in the pigmented basel cells of epidermal layer [10X Magnification].

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of TLR5 in formalin-fixed paraffin-embedded mouse brain tissue. Samples were incubated in TLR5 monoclonal antibody (Product # MA5-16237) using a dilution of 5 µg/mL.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Flow cytometry of TLR5 in CD14-/CD83+/TLR5+ diffferentiated myeloid dendritic cells (mDCs). Samples were incubated in TLR5 monoclonal antibody (Product # MA5-16237) using a dilution of 1 µg. Analysis using the PE conjugate of antibody. PBMCs were stimulated 5 days with GM-CSF+IL4 prior to the staining. The cells were stained for surface markers CD14 and CD83 FITC, and TLR5 PE. CD14- cells were gated and stained with PE-conjugated isotype control (mouse IgG2a), and FITC-conjugated CD83 (left). CD14- cells were gated and stained with PE-conjugated TLR5 , and FITC-conjugated CD83 (right).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Flow cytometry of TLR5 in human monocytes. Samples were incubated in TLR5 monoclonal antibody (Product # MA5-16237) using a dilution of 0.5 µg. Shaded histogram represents cells without antibody; green represents isotype control; red represents TLR5 antibody.

MA5-16237