Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [3]
- Immunocytochemistry [2]
- Immunohistochemistry [3]
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Validation data
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- Product number
- PA5-18248 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- VPS35 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- This antibody is predicted to react with bovine, mouse and rat based on sequence homology. This antibody is tested in Peptide ELISA: antibody detection limit dilution 128,000.
- Reactivity
- Human, Mouse, Rat
- Host
- Goat
- Isotype
- IgG
- Vial size
- 100 µg
- Concentration
- 0.5 mg/mL
- Storage
- -20° C, Avoid Freeze/Thaw Cycles
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western Blot staining of Human Cerebellum lysate using Product # PA5-18248 at a concentration of 0.1 µg/mL, the primary antibody incubation was 1 hour and the detection method was chemiluminescence.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- KD of VPS35 was achieved by transfecting HeLa with VPS35 specific siRNAs (Silencer® select Product # s31375, s31376). Western blot analysis (Fig. a) was performed using whole cell extracts from the VPS35 KD cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with VPS35 Polyclonal Antibody (Product # PA5-18248, 1µg/mL) and Rabbit Anti Goat IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27014, 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 VPS35.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-VPS35 Goat Polyclonal Antibody (Product # PA5-18248) and 80 kDa band corresponding to VPS35 was observed across the cell lines and tissues tested. Whole cell extracts (30 µg lysate) of U-87 MG (Lane1), A549 (Lane2), U-937 (Lane 3), SH-SY5Y (Lane 4), HeLa (Lane 5),tissue extracts of Mouse Lung (lane 6), Rat Lung (Lane 7) and Mouse Spleen (Lane 8) were electrophoresed using Novex® NuPAGE® 4-12% Bis-Tris gel (Product # NP0322BOX). 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 (1µg/mL) and detected by chemiluminescence Rabbit Anti-Goat IgG Secondary Antibody, HRP conjugate (Product # A27014, 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)
- Main image
- Experimental details
- Immunofluorescent analysis of VPS35 in HEK293 using a VPS35 polyclonal antibody (Product # PA5-18248). The sample was paraformaldehyde fixed, permeabilized with saponin, and detected with FITC in confocal microscopy. Data obtained from Aarhus University Denmark.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of VPS35 was performed using U-87 MG cells. 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 VPS35 Goat Polyclonal Antibody (Product # PA5-18248) at 5µg/mL in 0.1% BSA and incubated overnight at 4 degree and then labeled with Rabbit anti-Goat IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A-11078) 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 composite image showing endosomal localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemical analysis of VPS35 in Human Prostate using a VPS35 monoclonal antibody (Product #PA5-18248) at 5 µg/mL. The Human Prostate tissue section was paraffin embeded and detected using steamed antigen retrieval with citrate buffer pH 6, AP-staining.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemical staining of Epithelial cells of the epididymis (top) and Some leydig cells (bottom) using Product # PA5-18248 at a concentration of 3 µg/mL and 10 µg/mL respectively.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemical analysis of VPS35 in Human Small Intestine using a VPS35 monoclonal antibody (Product #PA5-18248) at 5 µg/mL. The Human Small Intestine tissue section was paraffin embeded and detected using steamed antigen retrieval with citrate buffer pH 6, AP-staining.