Journal: STAR Protocols

Article Title: Optimized protocol for direct cardiac reprogramming in mice using Ascl1 and Mef2c

doi: 10.1016/j.xpro.2023.102204

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Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse CD90.1 (Thy-1.1) Monoclonal Antibody (HIS51), Biotin(1:5) Thermo Fisher Cat# 13-0900-81 RRID AB_466530 Mouse Cardiac Troponin T Monoclonal Antibody (1:500) Thermo Fisher Cat# MA5-12960 RRID AB_11000742 Rabbit Recombinant Anti-Sarcomeric Alpha Actinin Antibody (1:500) Abcam Cat# ab68167 RRID AB_11157538 Alexa Fluor® 488 AffiniPure Donkey Anti-Mouse IgG (H+L) (1:500) Jackson ImmunoResearch Cat# 715-545-150 RRID AB_2340846 Alexa Fluor® 647 AffiniPure Donkey Anti-Rabbit IgG (H+L) (1:500) Jackson ImmunoResearch Cat# 711-605-152 RRID AB_2492288 Chemicals, peptides, and recombinant proteins Poly-l-lysine Electron Microscopy Sciences Cat# 1932-B DMEM with L-glutamine, 4.5 g/L glucose and sodium pyruvate Corning Cat# 10-013-CV IMDM Gibco Cat# 21980 Opti-MEM Gibco Cat# 31985070 FBS Millipore Sigma Cat# F2442-500ML 0.05% Trypsin Corning Cat# 25-052-CI M199 Corning Cat# 10-060-CV HBSS, 1× Corning Cat# 21-022-CV Collagenase II Worthington Cat# LS004177 2% Gelatin Millipore Sigma Cat# G1393 HyClone BSA Fraction V GE Cat# SH30574.02 0.5M EDTA Invitrogen Cat# 15575020 10 mg/mL Polybrene Millipore Cat# TR-1003-G Puromycin Thermo Fisher Cat# A1113803 Non-essential amino acids Gibco Cat# 25-025-CI Recombinant human VEGF165 (with carrier protein) R&D Cat# 293-VE Recombinant human FGF basic 146aa (with carrier protein) R&D Cat# 3718-FB Recombinant human FGF10 (with carrier protein) R&D Cat# 345-FG GlutaMAX Supplement Thermo Fisher Cat# 35050061 L-Ascorbic acid Millipore Sigma Cat# A4403 StemPro-34 SF medium Thermo Fisher Cat# 10639011 Critical commercial assays Trans IT®-LT1 Transfection Reagent Mirus Bio Cat# MIR 2305 Experimental models: Cell lines HEK293T ATCC Cat# CRL-3216 Experimental models: Organisms/strains CD-1 (mus musculus, CD1(ICR), P0-P3,Female and Male) Charles River Cat# 022 Recombinant DNA pMXs-puro-dsRed Wang et al. 2 N/A pCMV-VSV-G Addgene Cat# 8454 pUMVC Addgene Cat# 8449 pMXs-puro-Ascl1 Addgene Cat# 198227 pMXs-puro-Mef2c Addgene Cat# 198228 Other Open-Top Polypropylene Tube, 32mL Beckman Coulter 355642 MACS® MultiStand Miltenyi Biotec Cat# 130-042-303 MidiMACS™ Separator Miltenyi Biotec Cat# 130-042-302 LS columns Miltenyi Biotec Cat# 130-042-401 Anti-biotin microbeads Miltenyi Biotec Cat# 130-090-485 Open in a separate window .

Techniques: Recombinant, Electron Microscopy, Transfection