Alkaline Phosphatase Staining of piPSCs

The piPSCs (cultured for 5 days) were fixed in 4% paraformaldehyde (pH 7.4) at room temperature for 15 min, with AST Fast Red TR and α-naphthol AS MX phosphate (Sigma-Aldrich, USA) then used to stain the cells according to the manufacturer’s instructions. The piPSCs were incubated in 1.0 mg/mL Fast Red TR, 0.4 mg/mL α-naphthol AS-MX, and 0.1 mmol/L Tris-HCL 8.8 buffer at room temperature for 20 min. The AP-positive piPSCs clones showed red (Zhang et al., 2017 (link)). Images were obtained using a Nikon phase difference microscope.

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Wu X.L., Zhu Z.S., Xiao X., Zhou Z., Yu S., Shen Q.Y., Zhang J.Q., Yue W., Zhang R., He X., Peng S., Zhang S.Q., Li N., Liao M.Z, & Hua J.L. (2021). LIN28A inhibits DUSP family phosphatases and activates MAPK signaling pathway to maintain pluripotency in porcine induced pluripotent stem cells. Zoological Research, 42(3), 377-388.

Publication 2021

AST Fast Red TR α-naphthol AS MX phosphate

Clones Microscope Naphthol Naphthol as mx phosphate Paraformaldehyde Stain Tris buffer

Corresponding Organization :

Other organizations : Northwest A&F University

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Protocol cited in 3 other protocols

Variable analysis

independent variables

Culturing duration of piPSCs (5 days)

dependent variables

Alkaline phosphatase (AP) activity in piPSCs clones

control variables

Paraformaldehyde concentration (4%)
Paraformaldehyde pH (7.4)
Paraformaldehyde incubation time (15 minutes)
Staining reagents (AST Fast Red TR and α-naphthol AS MX phosphate)
Staining incubation time (20 minutes)
Staining buffer (0.1 mmol/L Tris-HCl, pH 8.8)
Imaging method (Nikon phase difference microscope)

controls

Positive control: AP-positive piPSCs clones showing red staining
Negative control: Not explicitly mentioned

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