PA5-28099

antibody from Invitrogen Antibodies

Targeting: SPHK1 SPHK

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Immunohistochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

PA5-28099

Provider

Invitrogen Antibodies

Product name

SPHK1 Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

Recommended positive controls: A431, Molt-4.

Concentration

1.17 mg/mL

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western Blot using SPHK1 Polyclonal Antibody (Product # PA5-28099). Sample (30 µg of whole cell lysate). Lane A: Molt-4. 10% SDS PAGE. SPHK1 Polyclonal Antibody (Product # PA5-28099) diluted at 1:500.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockdown of SPHK1 was achieved by transfecting Hep G2 with SPHK1 specific siRNAs (Silencer® select Product # S16959). Western blot analysis (Fig. a) was performed using Membrane Enriched lysate from the SPHK1 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with SPHK1 Polyclonal Antibody (Product # PA5-28099, 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 SPHK1.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-SPHK1 Polyclonal Antibody (Product # PA5-28099) and a 60 and 40 kDa bands corresponding to SPHK1 isoforms were observed across all cell lines tested. Membrane enriched lysate (40 µg lysate) of Hep G2 (Lane 1), HeLa (Lane 2), and U-87 MG (Lane 3) 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 with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant 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).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescent analysis of SPHK1 in paraformaldehyde-fixed A431 cells using a SPHK1 polyclonal antibody (Product # PA5-28099) at a 1:200 dilution.

Immunohistochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunohistochemical analysis of paraffin-embedded A549 xenograft, using SPHK1 (Product # PA5-28099) antibody at 1:100 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.