Module 4: Refining the Pluripotent Stem Cell Phenotype

hESC stained positively for DAPI (blue) and CD9 (green)

Module Leader Dr Andrew Laslett
Host Organisation CSIRO

Module description

The Laslett laboratory has developed a FACS-based immunotranscriptional profiling system for identifying and isolating hESCs that express high levels of the cell surface antigens CD9 and GCTM-2 by FACS (Laslett et al, BMC Developmental Biology 2007). This work has since been extended using multiple hESC lines and conditions and by combining immunotranscriptional and polysome translation state analyses (in collaboration with Dr Sean Grimmond, University of Queensland). The laboratory has used this technique to identify a list of stem cell markers (675 genes). Combining this information with 2614 genes identified by membrane polysome translation state analysis (MPTSA), the laboratory has identified  a large number of potential novel cell surface markers for hESC (Kolle et al, Stem Cells 2009).

Aims

This work is predicated on the hypothesis that combinations of cell surface markers can identify and isolate human embryonic stem cells (hESC) from heterogeneous cultures. Our previous work has identified a method to identify the cells within these heterogeneous cultures that are pluripotent and those that have taken the earliest steps of differentiation. In this module we will exploit this methodology to:

  1. identify and validate the genetic signatures of undifferentiated hESC and the earliest differentiated hESC progeny
  2. use these genetic signatures to identify, produce and characterise new cell surface markers for undifferentiated hESC.

    Module Leader biography

    Andrew and his team joined CSIRO Molecular Health Technologies in August 2009 and they remain based at the Australian Stem Cell Centre (ASCC) in Clayton. Prior to this he was a Senior Scientist and Group Leader of the Human Embryonic Stem Cell Technology Laboratory at the ASCC.  Dr Laslett’s research compares human embryonic stem cells to human induced pluripotent stem cells and is focused on exploiting the basic biology of these cell types to create novel tools that enhance pluripotent cell research translation. He leads an independent program as well as having significant national and international collaborations.

    In September 2007, Dr Laslett was elected as a Board Member and Director of the Australian Society of Medical Research (ASMR). Dr Laslett’s research is supported by the Australian Stem Cell Centre, the NHMRC, the Victoria California Stem Cell Alliance (Victorian Government and California Institute for Regenerative Medicine) and the NSW / Victorian Government Stem Cell Research Grant Program.

    Contact details

     E-mail  andrew.laslett@csiro.au 
     Phone  +61 3 9271 1133

    Selected publications

    1. Laslett AL, Filipczyk A & Pera MF (2003). Characterization and Culture of Human Embryonic Stem Cells. Trends In Cardiovascular Medicine 13(7):295-301.
    2. Martin F. Pera, Adam Filipczyk, Susan M. Hawes, and Andrew L. Laslett (2003) Isolation, characterization, and differentiation of human embryonic stem cells. Methods in Enzymology 365: 429-46.
    3. Martin F. Pera, Andrew Laslett, Susan M. Hawes, Irene Tellis, Karen Koh, and Linh Nguyen. (2006) Isolation and characterization of human embryonic stem cells.  In Embryonic stem cells:  a practical approach.  Edited by Elena Notarianni and Martin J. Evans. Oxford University Press, p238-259.
    4. Herszfeld D, Wolvetang E, Filipczyk A, Langton –Bunker E, Koh K, Nguyen L, Tellis I, Michalska A, Laslett AL, Reubinoff BE, Houssami S, Looijenga LH and Pera MF (2006) CD30 is a survival factor and a biomarker for transformed human pluripotent stem cells. Nature Biotechnology 24(3):351-7.  
    5. Andrew L. Laslett, Sean Grimmond, Brooke Gardiner, Lincon Stamp, Adelia Lin, Susan M. Hawes, Sam Wormald, David Nikolic-Paterson, David Haylock, and Martin F. Pera (2007)  Transcriptional Analysis of Early Lineage Commitment In Human Embryonic Stem Cells.  BMC Developmental Biology Mar 2;7:12.
    6. Adam A. Filipczyk, Andrew L. Laslett, Christine Mummery & Martin F. Pera. (2007) Differentiation is Coupled to Changes in the Cell Cycle Regulatory Apparatus of Human Embryonic Stem Cells. Stem Cell Research 1:1; 45-60.
    7. Andrew L. Laslett, Adelia Lin and Martin F. Pera (2007) Characterization and differentiation of human embryonic stem cells. In: Embryonic Stem Cells edited by .JR.Masters et al, Springer, p27–40.
    8. Andrew L. Laslett, Andrew Fryga and Martin F. Pera (2007) Flow cytometric analysis of human embryonic stem cells. In: Human Stem Cell Manual:  A Laboratory Guide, edited by Jeanne F. Loring, Elsevier, p96-107.
    9. Lina Gubhaju, Andrew Laslett, John F. Bertram, Anthony Zulli, Jane M Black (2008) Immunohistochemical localisation of TRA-1-60, TRA-1-81, GCTM-2 and podocalyxin in the developing baboon kidney. Histochemistry and Cell Biology 129(5):651-7.
    10. Kolle, G. Ho, SHM., Zhou, Q, Chy, H.S., Krishnan, K, Cloonan, N, Bertoncello, I, Laslett, A.L. and Grimmond, S.M.  Identification of human embryonic stem cell surface markers by combined membrane-polysome translation state array analysis and immunotranscriptional profiling. Stem Cells (in Press, July 2009)