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Western blot
ImmunoprecipitationAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [6]
- Immunocytochemistry [1]
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- Product number
- PA1-31840 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- NUMB Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- Recommended positive controls: The peptide used to generate this antibody is available for purchase (GTX24147-PEP).. Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.
- Reactivity
- Human, Mouse, Rat
- Host
- Goat
- Isotype
- IgG
- Vial size
- 50 µg
- Concentration
- 0.5 mg/mL
- Storage
- Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of NUMB using a NUMB polyclonal antibody (Product # PA1-31840).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of NUMB in A431 lysate (RIPA buffer, 30g total protein per lane) using a NUMB polyclonal antibody (Product # PA1-31840) at a dilution of 1g/mL following incubation for 1 hour and detected using chemiluminescence.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western Blot analysis of NUMB was performed by loading 35 µg (in RIPA buffer) of rat brain lysates. Proteins were transferred to a membrane and probed with a NUMB Polyclonal Antibody (Product # PA1-31840) at a dilution of 0.01 µg/mL.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western Blot analysis of NUMB was performed by loading 35 µg (in RIPA buffer) of rat brain lysates. Proteins were transferred to a membrane and probed with a NUMB Polyclonal Antibody (Product # PA1-31840) at a dilution of 0.01 µg/mL.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of NUMB was achieved by transfecting U-87 MG cells with NUMB specific siRNAs (Silencer® select Product # s16466, Product # s16465). Western blot analysis (Fig. a) was performed using whole cell extracts from the NUMB knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed using NUMB Polyclonal Antibody (Product # PA1-31840) and Rabbit anti-Goat IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27014). 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 NUMB isoform.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-NUMB Polyclonal Antibody (Product # PA1-31840) on whole cell extracts (30 µg lysate) of U-87 MG (Lane 1), PC-3 (Lane 2), DU 145 (Lane 3), A549(Lane 4), SH-SY5Y (Lane 5), HeLa (Lane 6), Mouse lung (Lane 7), Mouse brain (Lane 8) and Mouse Kidney (Lane 9) and numb major isoform 65, 53 kDa was observed across the samples tested. 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 (1:2000 dilution) and detected by Rabbit anti-Goat IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (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
- Immunofluorescence analysis of NUMB 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 1% BSA for 1 hour at room temperature. The cells were labeled with NUMB Polyclonal Antibody (Product # PA1-31840) at 5 µg/mL in 0.1% BSA, incubated at 4 degree Celsius overnight 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). Panel d represents the merged image showing Cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
