Anti cd24 antibody

For protein analysis, cells were harvested in SDS lysis buffer and sonicated. The determination of protein concentration was done using the Compat-Able Protein Assay Preparation Reagent Set (Thermo Scientific, #23215, Vienna, Austria) and the Pierce BCA Protein Assay Kit (Thermo Scientific, #23227, Vienna, Austria). Samples (20 µg total protein) were separated on a 10% SDS-polyacrylamide gel and blotted onto a PVDF membrane. The membranes were probed with anti-perilipin antibody (Cell Signaling, #9349, Frankfurt am Main, Germany), anti-PPARγ2 antibody (Cell Signaling, #2435, Frankfurt am Main, Germany), anti-FABP4 antibody (Cayman, #10004944, Tallinn, Estonia), anti-CD24 antibody (Abcam, #179821) and anti-GAPDH (Thermo Scientific, AM4300, Vienna, Austria). Goat anti-rabbit IgG-HRP (DAKO, Vienna, Austria) and goat anti-mouse IgG-HRP (Promega, Walldorf, Germany) served as secondary antibodies and signals were detected with a chemiluminescence detection system. Membranes were stained with Ponceau S for normalization to total protein. Densitometric analysis was done using ImageJ software.

Cell surface antigens for human iPSCs were analyzed with FACS. Cells (1 × 10⁵ cells per well) were incubated with one of the following primary antibodies: anti-CD24 antibody (Abcam), 1:100 dilution; anti-SSEA3 antibody (Abcam), 1:100 dilution; anti-CD105 antibody (Abcam), 1:200 dilution; anti-Nanog antibody (CST), 1:100 dilution; anti-Sox2 antibody (CST), 1:300 dilution; and anti-Oct4 antibody, 1:600 dilution (CST). After 1 h of incubation at 4 °C, the cells were washed, centrifuged, stained with 100 μl of a secondary antibody selected from anti-mouse IgG (H + L) Alexa Fluor® 488 Conjugate (CST) at 1:1000 dilution, anti-rat IgG (H + L) Alexa Fluor® 488 Conjugate (CST) at 1:1000 dilution, and anti-rabbit IgG (H + L) Alexa Fluor® 488 Conjugate (CST) at 1:1000 dilution. The stained cell pellets were suspended in 200 μl PBS/10 % FCS at 4 °C for flow cytometry analysis. All antibodies were validated for antigen specificity.