- PA5-64189 - Invitrogen Antibodies BHLHE22 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-BHLHB5 Polyclonal Antibody (Product # PA5-64189) and 37 kDa band corresponding to BHLHB5 was observed in Mouse Cerebellum in comparison to Mouse Kidney which is reported negative for BHLHB5 expression. An uncharacterized band was observed at ~50kDa in Mouse Cerebellum. Tissue extracts (30 µg lysate) of Mouse Cerebellum (Lane 1) and Mouse Kidney (Lane 2) were electrophoresed using Novex® 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 (1:500 dilution) and detected by chemiluminescence Goat Anti-Rabbit IgG Secondary Antibody, HRP conjugate (Product # A27036, 1:4000 dilution) 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: Immunofluorescence analysis of BHLHB5 was performed using log phase THP-1 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 BHLHB5 Polyclonal Antibody (Product # PA5-64189) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ 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 ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear localization. Panel e represents cells with no primary antibody to assess background. Panel f shows Jurkat cells with low expression of BHLHB5 as per gene expression databases. The images were captured at 60X magnification..

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of BHLHB5 in human testis using BHLHB5 Polyclonal Antibody (Product # PA5-64189).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of BHLHB5 in human liver using BHLHB5 Polyclonal Antibody (Product # PA5-64189).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of BHLHB5 in human colon using BHLHB5 Polyclonal Antibody (Product # PA5-64189).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of BHLHB5 in human cerebellum, colon, liver and testis using BHLHB5 Polyclonal Antibody (Product # PA5-64189) (A) shows similar protein distribution across tissues to an independent BHLHB5 Polyclonal Antibody (B).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of BHLHB5 in human seminal vesicle using BHLHB5 Polyclonal Antibody (Product # PA5-64189) shows moderate nuclear positivity in glandular cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of BHLHB5 in human skeletal muscle using BHLHB5 Polyclonal Antibody (Product # PA5-64189) shows no positivity in myocytes as expected.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical staining of BHLHB5 in human cerebellum using BHLHB5 Polyclonal Antibody (Product # PA5-64189) shows strong nuclear positivity in cells in molecular layer.

PA5-64189