- PA5-54183 - Invitrogen Antibodies ATXN2 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of Ataxin-2 was performed by loading 10 µg of WT (lane 1) and ATXN CRISPR KO (lane 2) HAP1 cell lysates in RIPA buffer onto a 4-15% gradient polyacrylamide gel. Proteins were transferred to nitrocellulose membrane and blocked in 5% milk. Ponceau stained transfer of blot is shown. Ataxin-2 was detected using an Ataxin-2 polyclonal antibody (Product # PA5-54183) at a dilution of 1:2,000 in 5% BSA in TBST overnight at 4 deg, followed by secondary antibody diluted to 0.2 µg/mL using Goat anti-Rabbit IgG (H+L) HRP antibody (Product # 65-6120). Chemiluminescent detection was performed using Pierce ECL Western Blotting Substrate (Product # 32106). Data courtesy of YCharOS Inc., an open science company with the mission of characterizing commercially available antibodies using knockout validation.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent staining of Ataxin 2 in human cell line U-2 OS shows positivity in cytoplasm. Samples were probed using an Ataxin 2 Polyclonal Antibody (Product # PA5-54183).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence of Ataxin 2 was performed using parental and Ataxin-2 HAP1 CRISPR KO cells that were transfected with a GFP or mCherry plasmid, respectively. At 48 hrs post transfection parental and KO cells were mixed and plated to a 1:1 ratio on coverslips as a mosaic and incubated for 24 hrs. Cells were fixed in 4% PFA for 15 min and permeabilized with 0.1% Triton X-100. Cells were stained with Ataxin polyclonal antibody (Product # PA5-54183) overnight at 4 deg. Secondary antibody incubation was performed using 1 µg/mL of goat anti-rabbit IgG (H+L) Alexa Fluor 555 secondary antibody (Product # A-21429) for 1 hr at RT. Imaging was performed with a 40X oil objective and analysis was performed using Image J. Cell image represents a single focal plane. Data courtesy of YCharOS Inc., an open science company with the mission of characterizing commercially available antibodies using knockout validation.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence of Ataxin 2 was performed using parental and Ataxin-2 HAP1 CRISPR KO cells that were transfected with a GFP or mCherry plasmid, respectively. At 48 hrs post transfection parental and KO cells were mixed and plated to a 1:1 ratio on coverslips as a mosaic and incubated for 24 hrs. Cells were fixed in 4% PFA for 15 min and permeabilized with 0.1% Triton X-100. Cells were stained with Ataxin polyclonal antibody (Product # PA5-54183) overnight at 4 deg. Secondary antibody incubation was performed using 1 µg/mL of goat anti-rabbit IgG (H+L) Alexa Fluor 555 secondary antibody (Product # A-21429) for 1 hr at RT. Imaging was performed with a 40X oil objective and analysis was performed using Image J. Cell image represents a single focal plane. Data courtesy of YCharOS Inc., an open science company with the mission of characterizing commercially available antibodies using knockout validation.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunoprecipitation of Ataxin was performed on HAP1 cell lysates. Antibody-bead conjugates were prepared by adding 1 µg of Ataxin polyclonal antibody (Product # PA5-54183) with 30 µL of protein A -Sepharose beads and rocked overnight at 4 deg. 1 µg of lysate was incubated with antibody-bead conjugate for 2 hrs at 4 deg. After multiple washes, 10% starting material (SM), 10% unbound fraction (UB) and immunoprecipitated fraction (IP) were processed for immunoblot using the same ATXN antibody (Product # PA5-54183). Ponceau stained transfer of blot is shown. Data courtesy of YCharOS Inc., an open science company with the mission of characterizing commercially available antibodies using knockout validation.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical staining of Ataxin 2 in human testis using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) shows moderate to strong cytoplasmic positivity in cells in seminiferous ducts.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of Ataxin 2 in human testis and skeletal muscle tissues using Ataxin 2 Polyclonal Antibody (Product # PA5-54183). Corresponding ATXN2 RNA-seq data are presented for the same tissues.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of Ataxin 2 in human colon, kidney, skeletal muscle and testis using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) (A) shows similar protein distribution across tissues to an independent Ataxin 2 Polyclonal Antibody (B).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical staining of Ataxin 2 in human cerebral cortex using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) shows moderate to strong cytoplasmic positivity in neurons.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of Ataxin 2 in human cerebellum using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) shows moderate to strong cytoplasmic positivity in purkinje cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical staining of Ataxin 2 in human skeletal muscle using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) shows weak to moderate cytoplasmic positivity in a subset of myocytes.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical staining of Ataxin 2 in human colon, kidney, skeletal muscle and testis using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) (A) shows similar protein distribution across tissues to an independent Ataxin 2 Polyclonal Antibody (B).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical staining of Ataxin 2 in human cerebral cortex using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) shows moderate to strong cytoplasmic positivity in neurons.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical staining of Ataxin 2 in human skeletal muscle using Ataxin 2 Polyclonal Antibody (Product # PA5-54183) shows weak to moderate cytoplasmic positivity in a subset of myocytes.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunoprecipitation of Ataxin was performed on HAP1 cell lysates. Antibody-bead conjugates were prepared by adding 1 µg of Ataxin polyclonal antibody (Product # PA5-54183) with 30 µL of protein A -Sepharose beads and rocked overnight at 4 deg. 1 µg of lysate was incubated with antibody-bead conjugate for 2 hrs at 4 deg. After multiple washes, 10% starting material (SM), 10% unbound fraction (UB) and immunoprecipitated fraction (IP) were processed for immunoblot using the same ATXN antibody (Product # PA5-54183). Ponceau stained transfer of blot is shown. Data courtesy of YCharOS Inc., an open science company with the mission of characterizing commercially available antibodies using knockout validation.

PA5-54183