Cite as: Cold Spring Harb. Protoc.; 2006; doi:10.1101/pdb.prot4403

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De Novo Isolation of Embryonic Stem (ES) Cell Lines from Blastocysts

Andras Nagy, Marina Gertsenstein, Kristina Vintersten, and Richard Behringer

This protocol was adapted from "Isolation and Culture of Blastocyst-Derived Stem Cell Lines," Chapter 8 of Manipulating the Mouse Embryo, 3rd edition, by Andras Nagy, Marina Gertsenstein, Kristina Vintersten, and Richard Behringer. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2003.

The first 100 words of the full text of this article appear below.


INTRODUCTION

The starting material for de novo isolation of stem cell lines can be either normal 3.5-days post coitum (dpc) expanded blastocysts or "delayed" blastocysts. Delayed blastocysts are usually collected 4-6 days after ovariectomy. For both groups of blastocysts, tissue culture procedures are similar. The only difference is the timing of the first disaggregation, because delayed blastocysts will initially grow more slowly.


MATERIALS

Reagents

recipe ES cell culture medium (ES-DMEM)

Expanded blastocysts (3.5 dpc) (see Collecting Blastocysts) or "delayed blastocysts" (see Ovariectomy for Induction of Blastocyst Implantation Delay)

Light paraffin oil (e.g., embryo-tested mineral oil from Sigma, or ES cell-qualified light mineral . . . [Full Text of this Article]

Equipment


METHOD


TROUBLESHOOTING


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