- PA5-57110 - Invitrogen Antibodies XRN1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using anti-XRN1 Polyclonal Antibody (Product # PA5-57110) and a 180 kDa band corresponding to XRN1 was observed across all the untreated cell lines tested and was induced in SH-SY5Y upon BDNF treatment. Modified whole cell extracts (1%SDS) (30 µg lysate) of U-2 OS (Lane 1), SH-SY5Y (Lane 2), SH-SY5Y treated with BDNF (100 ng/mL for 72 hr) (Lane 3), NIH/3T3 (Lane 4), PC-3 (Lane 5) and LNCaP (Lane 6) were electrophoresed using NuPAGE® 10 % Bis-Tris gel (Product # NP0302BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by wet transfer. The blot was probed with the primary antibody (1 µg/mL) and detected by chemiluminescence 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 SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent staining of XRN1 in human cell line U-2 OS shows positivity in plasma membrane & cytoplasm. Samples were probed using a XRN1 Polyclonal Antibody (Product # PA5-57110).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent staining of XRN1 in human cell line U-251 MG using a XRN1 Polyclonal Antibody (Product # PA5-57110) shows localization to plasma membrane and cytosol.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of XRN1 was performed using SH-SY5Y and SH-SY5Y treated with BDNF (100 ng/mL, 72 hours). The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with XRN1 Polyclonal Antibody (Product # PA5-57110) at 5 µg/mL concentration in 0.1% BSA and incubated overnight at 4 degree and then labeled with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, Alexa Fluor 488 (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 and e represents the merged image showing cytoplasmic localization. Expression of XRN1 was increased upon BDNF treatment in SH-SY5Y cells (Panel d) as compared to untreated SH-SY5Y cells (Panel e). 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: RNA Immunoprecipitation (RIP) assay of endogenous XRN1 protein using Anti-XRN1 Antibody: RIP assay was performed using Anti-XRN1 Recombinant Rabbit Polyclonal Antibody (Product # PA5-57110, 5 ug) on whole cell lysate from Hep G2 cells exposed to heat shock (45 degrees for 1 hour). Normal Rabbit IgG was used as a negative IP control. RNA purified by RiboPure™ RNA Purification Kit (Product # AM1924) was analyzed by RT-PCR using the Power SYBR® Green RNA-to-CT™ 1-Step Kit (Product # 4389986) with the primers pairs over IGF2, ACTB, MYC, RRP41, CCNA2, IGF2 mRNA and MALAT non-coding RNA. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

PA5-57110