MA5-25298

antibody from Invitrogen Antibodies

Targeting: SAMHD1 AGS5, HDDC1, Mg11, MOP-5, SBBI88

Flow cytometry

Antibody data

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Validation data

SAMHD1 protein structure - MA5-25298 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of SAMHD1 in 3T3 cells, mouse brain, mouse liver, mouse spleen lysate using 10 µg per lane. Samples were probed with SAMHD1 (Product # MA5-25298) monoclonal antibody at a dilution of 1:200.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of SAMHD1 in human tissue (1: Testis; 2: Omentum; 3: Uterus; 4: Breast; 5: Brain; 6: Liver; 7: Ovary; 8: Thyroid gland; 9: colon;10: spleen) samples using 10 µg per lane. Samples were separated by SDS-PAGE and probed with SAMHD1 (Product # MA5-25298) monoclonal antibody at a dilution of 1:200.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of SAMHD1 in HEK293T cells in untransfected (Left lane) and transfected (Right lane) samples using 5 µg per lane. The samples were separated by SDS-PAGE and probed with SAMHD1 (Product # MA5-25298) monoclonal antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of SAMHD1 in HepG2, HeLa, HT29, A549, COS7, Jurkat, MDCK, PC12, MCF7 cells using 35 µg per lane. Samples were probed with SAMHD1 (Product # MA5-25298) monoclonal antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-SAMHD1 Monoclonal Antibody (OTI1F6) (Product # MA5-25298) and a 72kDa band corresponding to SAMHD1 was observed in all cell and tissue lysates tested except Jurkat, U-937, Mouse Brain and Rat Brain [10.1016/s0165-2478(00)00276-5]. Modified whole cell lysates (1% SDS) (30 µg lysate) of THP-1 (Lane 1), Caco-2 (Lane 2), Jurkat (Lane 3), U-937 (Lane 4), Mouse Lung (Lane 5), Mouse Spleen (Lane 6), Rat Spleen (Lane 7), Mouse Brain (Lane 8) and Rat Brain (Lane 9) 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 (1:200 dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005). A ~25kDa band corresponding to tissue IgG was observed in some mouse tissue models.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of SAMHD1 in human CD4+T cells. Cells were probed with a IdU monoclonal antibody (Product # MA5-25298).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of SAMHD1 in human CD4+T cells. Cells were probed with a IdU monoclonal antibody (Product # MA5-25298).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of SAMHD1 in human CD4+T cells. Cells were probed with a IdU monoclonal antibody (Product # MA5-25298).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of SAMHD1 was performed using 70% confluent log phase Caco-2 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with SAMHD1 Monoclonal Antibody (OTI1F6) (Product # MA5-25298) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Donkey anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32766) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear and plasma 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)
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Experimental details: 1 FIGURE Automated image analysis-based quantification of SAMHD1 expression in the cohorts of the MCL Younger and MCL Elderly trial. A and B, Immunohistochemical staining of SAMHD1 (A) or CD3 (B) in one representative mantle cell lymphoma TMA core. Red arrows highlight CD3 + , T cell-rich areas. C, The histogram shows all cells of panel A by staining (DAB) intensity of SAMHD1. The two peaks represent the two different cell populations, as indicated. D, Primary B cell lymphoma lymph node-derived cells were stained for viability, CD3, CD19 and SAMHD1. Plot shows the SAMHD1 levels of B cells and T cells of the same lymph node (n = 8). The P value was calculated by the two-sided Wilcoxon test. E, A total of 378 TMA cores (two per patient) were stained for SAMHD1. The positivity was then evaluated by automated image analysis as described in panels A to C (see Section 2 for details) and by two independent pathologists with a ranking of 0, 1 or 2. Each dot represents the mean intensity of two cores (automated analysis, y axis) or the mean score of both pathologists (x axis). For each discrete level (x axis), additional box plots are depicted. DAB, diaminobenzidine [Color figure can be viewed at wileyonlinelibrary.com ]

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 4 FIGURE Chemotherapy combinations overcome the protective effect mediated by high SAMHD1 expression in B cell lymphoma cell lines. A, SAMHD1 expression of various B cell lymphoma cell lines (n = 16) was determined by western blot. B, Intensity values obtained by western blot were normalized and correlated with normalized median fluorescence intensity (MFI) obtained by intracellular flow cytometry. C and D, Cell lines shown in panel A were incubated with different chemotherapeutics as single compound (C) or in combination (D). C, Shown are the mean viabilities of four concentrations correlated with the corresponding normalized intensity of SAMHD1 western blot. D, The heatmaps illustrate the mean correlation coefficients ( R ) across all cell lines, as shown in panel A, but separated by each pair of concentrations. R and P values shown in panel B to C represent Pearson's correlation coefficient and corresponding P value [Color figure can be viewed at wileyonlinelibrary.com ]