Ab124765

The following antibodies are used in immunohistochemistry studies: anti-Lin28A (ab124765, Abcam, USA), anti-RAN (ab53775, Abcam, USA), anti- HSBP1 (ab83247, Abcam, USA), anti-Ki67 (D154094, BBI Life Sciences, China), anti-E-cadherin (20874-1- AP, Proteintech, China). The antibodies used for western blot analysis are as follows: anti RAN (10469- 1-AP, Proteintech, China), anti-HSBP1 (DF8954, Affinit, China), anti-Actin (AC004, ABclonal, China), anti-E-cadherin (20874-1-AP, Proteintech, China), anti-N-cadherin (22018-1-AP, Proteintech, China), anti-Vimentin (10366-1-AP, Proteintech, China), Apoptosis Antibody Sampler Kit (#9915T, CST, USA), Epithelial-Mesenchymal Transition (EMT) Antibody Sampler Kit (#9782, CST, USA).

The binding potential between LIN28A and GEFT/LINC00355 in transfected or un-transfected HCT-116 cells was measured through RIP assay at 48 h post transfection using the Magna RIP RNA-Binding Protein Immunoprecipitation Kit (Millipore, Temecula, CA, USA) together with IgG (ab172730, 1:20 dilution, Abcam) or LIN28A antibody (ab124765, 1:50 dilution, Abcam) according to the instructions of manufacturer. LINC00355 or GEFT mRNA level enriched by IgG or LIN28A antibody were determined by RT-qPCR assay.

Cells were lysed in Np40 buffer (Pierce, Appleton, WA, USA). Proteins were subjected to SDS-PAGE gel and transferred to a nitrocellulose membrane (GE Healthcare, Chicago, IL, USA). The membrane was blocked for 1 h in PBST containing 5% milk, and subsequently probed with primary antibody against LIN28 (Abcam, ab124765) overnight at 4 °C. After 1 h incubation with donkey-anti-rabbit HRP-conjugated secondary antibody (GE Healthcare, Chicago, IL, USA), the protein level was detected using SuperSignal West Femto Luminol reagents (Invitrogen, Waltham, MA, USA).