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Western blotAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [2]
- Immunocytochemistry [2]
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- Product number
- PA5-65272 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- GLB1 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Recombinant full-length protein
- Description
- Immunogen sequence: GRYWPARGPQL TLFVPQHILM TSAPNTITVL ELEWAPCSSD DPELCAVTFV DRPVIGSSVT YDHPSKPVEK RLMPPPPQKN KDSWLDHV
- Concentration
- 0.1 mg/mL
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of GLB1 was achieved by transfecting SK-BR-3 with GLB1 specific siRNAs (Silencer® select Product # s5797). Western blot analysis (Fig. a) was performed using Whole cell extracts from the GLB1 knockdown cells (lane 2) and non-targeting scrambled siRNA transfected cells (lane 1). The blot was probed with GLB1 Polyclonal Antibody (Product # PA5-65272, 1:500 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 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 GLB1.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-GLB1 Polyclonal Antibody(Product # PA5-65272) and a 76kDa band corresponding to GLB1 was observed across cell lines tested. Whole cell extracts (30 µg lysate) of BeWo (Lane 1), Daudi (Lane 2), U-87 MG (Lane 3), A549 (Lane 4), HeLa (Lane 5) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). 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 (1:500 dilution) and detected by chemiluminescence with 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 Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005). An uncharacterized band was also observed at ~120kDa.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent staining of beta Galactosidase in human cell line A-431 shows localization to the Golgi apparatus and vesicles. Samples were probed using a beta Galactosidase Polyclonal Antibody (Product # PA5-65272).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of GLB1 was performed using 70% confluent log phase U-87 MG 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 45 minutes at room temperature. The cells were labeled with GLB1 Polyclonal Antibody (Product # PA5-65272) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790), (1:2000 dilution), 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 cytoplasmic (golgi, ER pattern) localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
