Class Syllabus
Day One:
9 a.m. - 10 a.m. Introduction to general hES cell culture techniques and work plan on lab sessions
10 a.m. - noon Lab Session
- QC Regent preparation
- Observation of Feeder and hES cell cultures
- Start Derivation of MEF
Noon - 1 p.m. Lunch and informal talk "How to QC hES cells"
2 p.m. - 4 p.m. Lab session
- MEF derivation
- Thawing and plating Feeders
- Thawing and patting hES cells
Day Two:
9 a.m. - noon Lab Session
- Observation of Feeder and hES cell cultures that were plated on Monday
- Expanding MEF we will discuss how to do the expansion at your lab
(After Mycoplasma testing you will expand, irradiate and freeze in your own lab)
Noon - 1 p.m. Lunch and informal talk about “Planning differentiation experiments with hES cells”
1 p.m. - 4 p.m. Lab Session
- Review hES cell morphology
- Identify differentiated colonies
- Remove differentiated colonies using colony marker and stereoscope
- Passaging hES cell enzimatically, manually or picking specific colonies
- Freezing hES cell
Day Three:
- Review hES cell morphology
- QC Immunohistochemistry protocol Oct4 and SSEA4
- Freezing MEF p1 to continue expansion at your lab
Noon - 1p.m. Lunch and review protocol, questions and answers
1 p.m. - 4 p.m. QC
- Test pluripotency by Teratoma formation
- FACS analysis for OCT4 and SEEA4 and SSEA1
- Karyotyping
- aCGH
- Mycoplasma