Dec 19, 2023

Public workspaceTrophoblast Organoid Media (TOM)

This protocol is a draft, published without a DOI.
  • 1Duke University
vfarmer Farmer
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Protocol Citation: vfarmer Farmer 2023. Trophoblast Organoid Media (TOM). protocols.io https://protocols.io/view/trophoblast-organoid-media-tom-c6e9zbh6
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: December 15, 2023
Last Modified: December 19, 2023
Protocol Integer ID: 92353
Abstract
Media is also used with flipped TOs. Media can be stored at 4C and used within 1 month.
Recipe
Recipe
In a 50 mL conical add 40 mL of F12/DMEM. Mix in the following components:
B27 Amount1 mL

stock is 10X, final is 1X

supplier: ThermoFisher (Gibco), cat#: 17504044
N-Acetyl-L-Cystine (NAC) Amount625 µL

stock is 100 mM in H20, final is 1.25 mM

supplier: Sigma, cat#: A9165

N2 Amount500 µL

stock is 10X, final is 1X

supplier: ThermoFisher (Gibco), cat#: 17502048
L-Glutamine Amount500 µL

stock is 200 mM, final is 200 mM

supplier: ThermoFisher (Gibco), cat#: A2916801
Nicotinamide (NTM) Amount500 µL

stock is 1 M in H20, final is 10 mM

supplier: Sigma-Aldrich, cat#: N0636
Primocin Amount100 µL

stock is 50 mg/ml, final is 100 µg/ml

supplier: Inviogen, cat #: NC9141851
Y-27632 Amount100 µL

stock is 2.5 mM in H20, final is 5 µM
hEGF Amount25 µL

stock is 100 µg/ml in H20, final is 50 ng/ml

supplier: Peprotech, cat#: AF-100-15
hFGF2 Amount25 µL

stock is 200 µg/ml in 5mM Tris pH 7.6, final is 100 µg/ml

supplier: Peprotech, cat#: 100-18C
hHGF Amount25 µL

stock is 100 µg/ml in H20, final is 50 ng/ml

supplier: Peprotech, cat#: 100-39
R-spondin1 Amount25 µL

stock is 160 µg/ml in PBS+0.1% BSA, final is 80 µg/ml

supplier: Bio-techne, cat#: 4645-RS
PGE2 Amount25 µL

stock is 5 mM in acetone, final is 2.5 µM

supplier: Sigma, cat#: P0409
A83-01 Amount5 µL

stock is 5 mM in DMSO, final is 500 nM

supplier: Tocris, cat#: 2939
CHIR99021 Amount5 µL

stock is 15 mM in DMSO, final is 1.5 µl

supplier: Tocris, cat#: 4423
FBS 5 mL

supplier: Cytiva, cat# SH30088.03HI