Antibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [3]
- ELISA [3]
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- Product number
- LS-C18846 - Provider product page
- Provider
- LSBio
- Product name
- MAP3K5 / ASK1 Antibody (phospho-Ser83) LS-C18846
- Antibody type
- Polyclonal
- Description
- Delipidation, salt fractionation and ion exchange chromatography followed by dialysis.
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Storage
- Store vial at -20°C prior to opening. Centrifuge product if not completely clear after standing at room temperature. Dilute only prior to immediate use. For extended storage aliquot contents and freeze at -20°C or below.
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Supportive validation
- Submitted by
- LSBio (provider)
- Enhanced method
- Genetic validation
- Main image
- Experimental details
- Anti-ASK-1 phospho specific pS83 Antibody - Western Blot. Western blot of anti-pS83 ASK1 antibodies shows specificity for phosphorylated human ASK1. Anti-pS83 (aa 76-87) antibody was tested by western blot against Cos-7 cell lysates after transient transfection with 1) vector only, 2) recombinant HA-ASK1, and 3) recombinant human HA-ASK1 where S83 was substituted with an alanine residue. Cells were lysed 24 h post-transfection in 200 ul of 1x SDS-sample buffer, heated at 96?C for 5, and vortexed for 30 sec. Samples (10 ul each) were separated on a 12% SDS-PAGE gel and transferred to PVDF (Millipore) followed by blocking for 45 using TTBS supplemented with 5% non-fat dry milk. All incubations were performed at room temperature. In panel a) all samples were incubated with anti-HA antibody. This blot demonstrates both recombinant transfections express rASK1. In panel b) all samples were incubated with anti-pS83 ASK1. Lane 2 shows strong specific staining of ASK1. Lane 3, where S83 was replaced with alanine, shows greatly diminished staining. In panel c) all samples were incubated with anti-pS83 ASK1 antibody as before except the antibody was pre-incubated with phospho peptide prior to membrane incubation. No staining is observed after phospho peptide blocking occurs.
- Submitted by
- LSBio (provider)
- Enhanced method
- Genetic validation
- Main image
- Experimental details
- Anti-ASK-1 phospho specific pS83 Antibody - Western Blot. Western blot of anti-pS83 ASK1 antibodies shows specificity for phosphorylated human ASK1. Anti-pS83 (aa 76-87) antibody was tested by western blot against Cos-7 cell lysates after transient transfection with 1) vector only, 2) recombinant HA-ASK1, and 3) recombinant human HA-ASK1 where S83 was substituted with an alanine residue. Cells were lysed 24 h post-transfection in 200 ul of 1x SDS-sample buffer, heated at 96?C for 5, and vortexed for 30 sec. Samples (10 ul each) were separated on a 12% SDS-PAGE gel and transferred to PVDF (Millipore) followed by blocking for 45 using TTBS supplemented with 5% non-fat dry milk. All incubations were performed at room temperature. In panel a) all samples were incubated with anti-HA antibody. This blot demonstrates both recombinant transfections express rASK1. In panel b) all samples were incubated with anti-pS83 ASK1. Lane 2 shows strong specific staining of ASK1. Lane 3, where S83 was replaced with alanine, shows greatly diminished staining. In panel c) all samples were incubated with anti-pS83 ASK1 antibody as before except the antibody was pre-incubated with phospho peptide prior to membrane incubation. No staining is observed after phospho peptide blocking occurs.
- Submitted by
- LSBio (provider)
- Enhanced method
- Genetic validation
- Main image
- Experimental details
- Anti-ASK-1 phospho specific pS83 Antibody - Western Blot. Western blot of anti-pS83 ASK1 antibodies shows specificity for phosphorylated human ASK1. Anti-pS83 (aa 76-87) antibody was tested by western blot against Cos-7 cell lysates after transient transfection with 1) vector only, 2) recombinant HA-ASK1, and 3) recombinant human HA-ASK1 where S83 was substituted with an alanine residue. Cells were lysed 24 h post-transfection in 200 ul of 1x SDS-sample buffer, heated at 96?C for 5, and vortexed for 30 sec. Samples (10 ul each) were separated on a 12% SDS-PAGE gel and transferred to PVDF (Millipore) followed by blocking for 45 using TTBS supplemented with 5% non-fat dry milk. All incubations were performed at room temperature. In panel a) all samples were incubated with anti-HA antibody. This blot demonstrates both recombinant transfections express rASK1. In panel b) all samples were incubated with anti-pS83 ASK1. Lane 2 shows strong specific staining of ASK1. Lane 3, where S83 was replaced with alanine, shows greatly diminished staining. In panel c) all samples were incubated with anti-pS83 ASK1 antibody as before except the antibody was pre-incubated with phospho peptide prior to membrane incubation. No staining is observed after phospho peptide blocking occurs.
Supportive validation
- Submitted by
- LSBio (provider)
- Enhanced method
- Genetic validation
- Main image
- Experimental details
- Anti-ASK-1 phospho specific pS83 Antibody - ELISA. ELISA results of purified polyclonal anti-pS83 ASK-1 (aa 76-87) antibody tested against BSA conjugates of non-phospho and phospho forms of immunizing peptide. Each well was coated with 0.1 mg of conjugate. The starting dilution of antibody was 1:1000 and each point on the X-axis represents a 2-fold dilution. HRP conjugated Gt-a-Rabbit IgG H&L and TMB substrate were used for detection.
- Submitted by
- LSBio (provider)
- Main image
- Experimental details
- Anti-ASK-1 phospho specific pS83 Antibody - ELISA. ELISA results of purified polyclonal anti-pS83 ASK-1 (aa 76-87) antibody tested against BSA conjugates of non-phospho and phospho forms of immunizing peptide. Each well was coated with 0.1 mg of conjugate. The starting dilution of antibody was 1:1000 and each point on the X-axis represents a 2-fold dilution. HRP conjugated Gt-a-Rabbit IgG H&L and TMB substrate were used for detection.
- Submitted by
- LSBio (provider)
- Main image
- Experimental details
- Anti-ASK-1 phospho specific pS83 Antibody - ELISA. ELISA results of purified polyclonal anti-pS83 ASK-1 (aa 76-87) antibody tested against BSA conjugates of non-phospho and phospho forms of immunizing peptide. Each well was coated with 0.1 mg of conjugate. The starting dilution of antibody was 1:1000 and each point on the X-axis represents a 2-fold dilution. HRP conjugated Gt-a-Rabbit IgG H&L and TMB substrate were used for detection.