Stem Cell Protocols

The combination of prospective identification/isolation of bone marrow progenitors and quantitative RT-PCR is a powerful tool to understand the molecular mechanism underlying hematopoiesis. Here, we described our standard procedures of the murine myeloid progenitor staining for fluorescence activated cells sorting (FACS) and RNA purification methods. - [Read Cell Staining for Sorting of Hematopoietic Stem Cells and Myeloid Progenitors and Isolating RNA]

This protocol decribes derivation of TS cell lines from 3.5-days post coitum (dpc) mouse blastocysts. The procedure is similar to the derivation of embryonic stem (ES) cell lines. However, the success rate is considerably higher, and less expertise is required to recognize pluripotent TS cell colonies. - [Read Derivation of Trophoblast Stem (TS) Cell Lines from Blastocysts Protocol]

Pluripotent ES cells can develop into many types of differentiated tissues if they are placed back into a differentiating environment. Often, differentiation proceeds through an intermediate stage called the embryoid body (EB). EBs can be manipulated further to generate more differentiated cell types. This protocol describes a method for differentiation of ES cells into EBs. - [Read Differentiating Embryonic Stem (ES) Cells into Embryoid Bodies Protocol]

Protocol for the electroporation of ES cells. Cells are routinely passaged two days prior to electroporating. Usually one 10 cm plate at approximately 80% confluency will provide enough cells for 1-2 electroporations. - [Read Electroporation of ES Cells Protocol]

This protocol describes a method for freezing and thawing ES cells using cryovials. It is important to freeze ES cell stocks as soon as possible to reduce the time that they are in culture. A careful record should be kept of the number of times cells are passaged and the location of the cryovials. - [Read Freezing and Thawing of Embryonic Stem (ES) Cells Using Cryovials Protocol]

This protocol describes passage of ES cells. They should be split at 1:3 to 1:7 every 2-3 days depending on their growth rate when they reach 70% confluency. They should never be allowed to grow past 90% confluency, but rather they should form tightly packed colonies not touching each other. - [Read Passage of Embryonic Stem (ES) Cells Protocol]

This protocol describes a method of DNA preparation from ES cells in 24-well plates. - [Read Preparation of DNA from Cells in 24-Well Tissue Culture Plates Protocol]

Ths protocol describes the preparation of ES cells, which can then be aggregated with diploid or tetraploid embryos to produce aggregation chimeras. - [Read Preparing Embryonic Stem (ES) Cells for Aggregation Protocol]

This protocol describes a method for rapid preparation of DNA from cells in 96-well tissue culture dishes. - [Read Rapid Preparation of DNA from Cells in 96-Well Tissue Culture Dishes Protocol]

Picking Transfected ES Cell Clones Protocol

This protocol a protocol on how to generate transfected embryonic stem (ES) cell clones. The previous protocol in this series is the Protocol for Electroporation of ES cells. The next protocol in the series is the Protocol on Disaggregation, Expansion, and Freezing of Transfected ES Clones.

[Read more]

Microinjection Pipette Pulling Protocol

A simple protocol for microinjection pipette pulling.

[Read more]

Acid Washing Microinjection Pipettes Protocol

Acid Washing of Glass microinjection pipettes protocol.

[Read more]

Microinjection Pipettes Protocol

Microinjection Pipettes Preparation Protocol.

[Read more]