Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [4]
- Immunocytochemistry [2]
- Immunohistochemistry [4]
- Flow cytometry [1]
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Validation data
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- Product number
- PA5-79587 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- LCK Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- Reconstitute with 0.2 mL of distilled water to yield a concentration of 500 µg/mL.
- Reactivity
- Human, Mouse, Rat
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µg
- Concentration
- 500 µg/mL
- Storage
- -20°C
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of LCK in Lane 1: HUT whole cell lysate, Lane 2: JURKAT whole cell lysate, Lane 3: RAJI whole cell lysate, Lane 4: CEM whole cell lysate, Lane 5: K562 whole cell lysate using 40-50 µg per well. Sample was incubated with LCK (Product # PA5-79587) at a dilution of 0.5 µg/mL.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of Lck in, Lane 1: human Jurkat whole cell lysates, Lane 2: mouse spleen tissue lysates, Lane 3: mouse thymus tissue lysates. Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 µg of sample under reducing conditions. After Electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. The membrane was blocked with 5% non-fat milk/TBS for 1. 5 hour at RT. The membrane was incubated with LCK Polyclonal Antibody (Product # PA5-79587) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0. 1% Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5,000 for 1. 5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit. A specific band was detected for Lck at approximately 58 kDa. The expected band size for Lck is at 58 kDa.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of LCK was achieved by transfecting Jurkat with LCK specific siRNAs (Silencer® select Product # s8108, s8107). Western blot analysis (Fig. a) was performed using whole cell extracts from the LCK knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with LCK Polyclonal Antibody (Product # PA5-79587, 1:1000 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 LCK.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-LCK Polyclonal Antibody (Product # PA5-79587) and a 54kDa band corresponding to LCK was observed in Jurkat, MOLT-4 and A549, but not in Ramos, SK-BR-3 and SK-OV-3 which are reported to be negative. Whole cell lysates (30ug lysate) of Jurkat (Lane 1), Ramos (Lane 2), MOLT-4 (Lane 3), A549 (Lane 4), SK-BR-3 (Lane 5) and SK-OV-3 (Lane 6) 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 bot was probed with the primary antibody (1:1000 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).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemistry analysis of Lck using anti-Lck antibody (Product # PA5-79587). Lck was detected in a section of U2OS cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent for 15 mins. The cells were blocked with 10% goat serum and then incubated with 2μg/mL rabbit anti-Lck antibody (Product # PA5-79587) overnight at 4°C. DyLight®594 Conjugated Goat Anti-Rabbit IgG was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.
- Submitted by
- Invitrogen Antibodies (provider)
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- Experimental details
- Immunofluorescence analysis of LCK was performed using Jurkat 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 LCK Polyclonal Antibody (Product # PA5-79587) at 5µg/mL in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 (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 membrane 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
- Immunohistochemistry analysis of LCK on paraffin-embedded rat Lymphaden tissue. Sample was incubated with LCK polyclonal antibody (Product# PA5-79587).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry analysis of LCK on paraffin-embedded human tonsil tissue. Sample was incubated with LCK polyclonal antibody (Product# PA5-79587).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry analysis of LCK on paraffin-embedded mouse Lymphaden tissue. Sample was incubated with LCK polyclonal antibody (Product# PA5-79587).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemical analysis of Lck in frozen section of mouse spleen tissue. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/mL rabbit anti-Lck antibody (Product # PA5-79587) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Flow Cytometry of LCK in HepG2 cells (blue line), isotype control rabbit IgG (green line) and unlabeled (red line). Samples were blocked with 10% goat serum, incubated with LCK Polyclonal Antibody (Product # PA5-79587) at a dilution of 1 μg (per 1x10^6 cells), followed by DyLight®488 conjugated goat anti-rabbit IgG (for 30 minutes at 20°C) using 5-10 μg (per 1x10^6 cells) dilution.