This is a knockout-validated antibody summary, based on the publication "Twist1-induced dissemination preserves epithelial identity and requires E-cadherin", as cited below [1]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

Figure 1. Western blot of organoids isolated from a Cre-ER;E-cadfl/fl mouse after E-cad deletion induced with tamoxifen (Tam). N-cadherin serves as loading control. From [1].

Rat monoclonal IgG2a

Company: Invitrogen

Antibody: E-cad

Catalog number: 13-1900

Summary: Rat monoclonal IgG2a against a mouse liver E-cadherin fragment. It reacts against human, mouse , and dog. Recommended for western blot, immunofluorescence, immunohistochemistry, flow cytometry and immunoprecipitation.

Figure 2. Immunofluorescence of E-cad in genetic mosaic E-cadfl/fl;mT/mG mammary ductal outgrowths. Cre+ (green) cells lack E-cad (arrowheads in D and F). Bars, 10 µm. From [1].

Western blot | Immunofluorescence

Fragments of primary mammary epithelial ducts, termed organoids, from Cre-ER;E-cadfl/fl;mT/mG mice 8–12 wk old.

WB: Lysis buffer for protein extraction was prepared by diluting 10× RIPA buffer (20–188; EMD Millipore) in ultrapure water and chilling the mixture at 4°C for at least 2 h. Immediately before use, lysis buffer was supplemented with 0.1% SDS, 5% glycerol, 3 mM EDTA, 1 mM NaF, 1 mM PMSF, 1.5 mM NaVO4, Aprotinin (A6279; Sigma-Aldrich), and a mini protease inhibitor tablet (11836153001; Roche).

IF: Organoids grown in 3D Matrigel were fixed in 4% paraformaldehyde and embedded in OCT and cut with a cryostat.

WB: 5% milk in TBST (03-500-537; Thermo Fisher Scientific) for 1 h at room temperature.

IF: 10% FBS/1% BSA for 1–3 h.

WB: 1:1,000 in blocking buffer overnight at 4°C.

IF: 1:250 in 1% FBS/1% BSA overnight at 4°C.

WB: HRP-conjugated secondary antibodies (1:2,000; Invitrogen) in blocking buffer for 1 h at room temperature.

IF: Alexa Fluor–conjugated antibodies (1:200; Invitrogen) in 1% FBS/1% BSA overnight at 4°C or for 2 h at room temperature.

WB: ECL reagents (34075 or 34095; Thermo Fisher Scientific), and membranes were imaged using an imager and software (Alpha-InnoTec).

IF: spinning-disk confocal microscope (Solamere Technology Group Inc.) with an XR/MEGA-10 S30 camera (Stanford Photonics, Inc.).

The same clone (ECCD-2) is sold as LifeSpan Biosciences LS-C12335; Enzo Life Sciences ALX-804-202-C100; Invitrogen 13-1900.