Journal: Nature Protocols

Article Title: Directed differentiation of human induced pluripotent stem cells into mature kidney podocytes and establishment of a Glomerulus Chip

doi: 10.1038/s41596-018-0007-8

Figure Lengend Snippet: Fig. 3 | Immunofluorescence staining and scanning electron micrograph of cells during the differentiation process. a–c, Human iPS cell–derived mesoderm cells expressing HAND1, goosecoid, and brachyury (a), intermediate mesoderm cells expressing WT1 and Pax2 markers (b), and podocytes expressing the lineage-characterization markers nephrin, WT1 and podocin, as well as the associated protein ApoL1 (c). d, High-magnification image of iPS cell–derived podocytes showing the development of foot processes that are positive for podocin. e, Scanning electron micrograph of human iPS cell–derived podocytes showing the development of primary and secondary foot processes. Scale bars, (a–c) 100 µm, (d) 25 µm, and (e) 2 µm. d,e, adapted with permission from Musah et al.5, Macmillan Publishers Limited.

Article Snippet: ● All-trans retinoic acid (500 mg; Stem Cell Technologies, cat. no. 72262) ● Heat-inactivated FBS (Thermo/Life Technologies, cat. no. 10082147) ● Penicillin–streptomycin, liquid (100×; Thermo/Life Technologies, cat. no. 15140-163) Antibodies and fluorescent stains ● Oct4 (R&D Systems, cat. no. AF1759) ● TRA-1-60 (Abcam, cat. no. ab16288) ● Goosecoid (R&D Systems, cat. no. AF4086) ● Brachyury (Abcam, cat. no. ab20680) ● HAND1 (Abcam, cat. no. ab196622) ● Pax2 (Invitrogen, cat. no. 71-6000) ● OSR1 (Novus Biologicals, cat. no. H00130497-M04 ● WT1 (Millipore, cat. no. MAB4234) ● Nephrin (Progen, cat. no. GP-N2) ● Podocin (Abcam, cat. no. ab50339) ● VE-cadherin (Santa Cruz Biotech, cat. no. sc-9989) ● Collagen type IV (Abcam, cat. no. ab6586) ● Integrin β1 (Abcam, cat. no. ab24693) ● ApoL1, anti-apolipoprotein L1 (Abcam, cat. no. ab85795) ● 4′,6-Diamidino-2-phenylindole (DAPI; Thermo/Life Technologies, cat. no. D1306) ● Oct4 conjugated to phycoerythrin (Oct4-PE; Stem Cell Technologies, cat. no. 60093PE) ● Alexa Fluor 488– and Alexa Fluor 594–conjugated secondary antibodies (Thermo/Life Technologies) ● WTI conjugated to APC (WT1-APC; LifeSpan Biosciences, cat. no. LS-C224662/5744) ● Nephrin conjugated to PE-Cy5 (Nephrin-PE-Cy5; Bioss, cat. no. bs-10233R-PE-Cy5) ECM molecules ● Matrigel hESC-qualified matric, 5-mL vial (BD Biosciences, cat. no. 354277).

Techniques: Staining, Derivative Assay, Expressing