This is a knockout-validated antibody summary, based on the publication cited below [1]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

Nbs1, also called AT-V1; AT-V2; ATV; NBS; NBS1; P95; Nijmegen breakage syndrome 1 (nibrin); cell cycle regulatory protein p95; p95 protein of the MRE11/RAD50 complex.

Figure 1. Novus. Western blotting of NBS1 levels from embryonic fibroblast cultures derived from E14.5 embryos. W: wild type; E: Exo1-KO; N: Nbs1-KO; NE: Exo- Nbs1-KO. From Figure 1 [1].

NB100-143 from Novus Bio, now part of Biotechne.

A301-284A from Bethyl, was used only in one experiment.

Novus: Rabbit polyclonal antibody that detects human, mouse, hamster, mammal and primate. Has been used in WB, ChIP, Flow, ICC/IF, IHC, IP

Bethyl: rabbit polyclonal antibody detecting mouse protein. Immunogen peptide was between aa 701 and 751, has been successfully used in western blot and immunoprecipitation. Predicted to recognise rat, bovine, dog and horse proteins.

Figure 2. Bethyl. Western blotting of cells transfected with or without a vector expressing human NBS1. W: wild type; N: Nbs1-KO; NE: Exo- Nbs1-KO. From Figure 4 [1].

Novus and Bethyl

Western blot

Primary MEFs extracted with NETN (20mM Tris, pH 8, 100 mM NaCl, 1 mM EDTA, 0.5% IGEPAL, 1.25 mM DTT, and Roche Protease Inhibitor Mixture) using several freeze/thaw cycles.

In Fig 1, the authors used Novus antibody NB100-143 (1:1000). Cells were lysed TNG-150 lysis buffer containing 50 mM Tris–HCl, 150mM NaCl, 1% Tween 20, 0.5% NP-40, 1× Phosphatase Inhibitor Cocktail 2 (Sigma–Aldrich) and 1× Protease Inhibitor Cocktail (Roche). Protein concentration was quantified with DC Protein Assay Kit (Bio-Rad). 30–80 μg of total protein was separated by SDS-PAGE (8–12% gels) or NuPAGE Novex Tris-Acetate SDS (3–8% gels, Life Technologies) followed by transfer to Immobilon-P PVDF membrane (Merck-Millipore). For blocking solution, 5% milk in PBST was used, membranes were incubated in primary antibody at 4°C overnight in secondary antibody 1 h at room temperature. ECL reagent (Amersham) with X-ray film (Fujifilm) were used to detect the signal.