Recently i extract human stem cells from dental pulp. How can i confirm that these cells are stem cells and aren't fibroblast.
I will perform flow cytometry for surface CD markers relatedto SCs. Can i detect SCs morphologically?
Dear Saeid,
Morphologically you really cannot distinguish them, its best to do flow sorting.
You can also do qualitative tests such as immunofluoresence.
Regards,
Prasad.
hi
In the case of embryonic stem cells and spermatogonial stem cells they usually grow in spherical and colony-like shape in culture flask , but I have no experience in this area. The best marker for the stem ness confirming as OCT4-A .
I hope this file will help you.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4478630/pdf/WJSC-7-839.pdf
Be happy & Good Luck
How about immunostaining your cells with the antibodies against the stem cell markers Oct4, Lin28, and Sox2?
Hello
You can use Fluorescence-activated cell sorting , Immunohistochemistry , Immunocytochemistry .
For specific markers for Dental pulp cells have been well described by researchers such as Dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP1) are the markers of odontoblast differentiation , STRO 1 and CD271 also , you can use MSCs surface markers and identify the expression rate where it owns different expression rates . you can refer to Misako Nakashima and et al where use a surface markers of MSCs ( positive marker : such as CD29, CD44, CD73, CD90, CD105, CD166, and STRO-1; and negative markers such as : CD11b, CD19, CD34, CD45, and HLA-DR ) but the expression rate of CXCR4, GCSFR, and CD105 in MDPSCs or fractionated DPSCs such as CD31- SP cells and CD105+ cells is significantly higher compared with unfractionated clonogenic DPSCs and the expression of Sox2 and CXCR4 mRNA in human MDPSCs and pulp CD105+ cells is lower compared with iPS cells, other stem cell .
Good Luck
http://cvgl.stanford.edu/teaching/cs231a_winter1415/prev/projects/embroynic-stem-cells.pdf
https://www.ncbi.nlm.nih.gov/pubmed/25173163
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4478630/
by using immunocytochemestry. there are many CD markers to detect the cells positive and negative
First you will need to verify what you have, that can be accomplished by Asymmetrex (contact information below). Then you can run immunofluorescence with or without flow cytometry to define cell surface markers for future reference. By the way, the true MSCs defined by Caplan, were a tripotent progenitor cell that could only form fat, cartilage, and bone. If you are looking for a pluripotent stem cell, be sure you have not isolated a mixture of cells, each with a few capabilities, but in toto demonstrate presumed pluripotency. That is where the Asymmetrex system comes in handy. Contact information:
James L. Sherley, M.D., Ph.D.
Director
Asymmetrex, LLC
P.O. Box 301179
Boston, MA 02130
USA
Phone: (617) 990-6819
Website http://asymmetrex.com
Facebook https://www.facebook.com/asymmetrex
Twitter https://twitter.com/asymmetrex
Saeid,
I agree with Saleh Alkarim and Henry Young. The CD markers for stem cell lineage are quite well defined, both for murine and for human cells. Some variation between species do exist. Be very careful considering markers such as OCT4, SOX2, etc, because their expression is relevant in embryonic stem cells, and teeth are not present in the embryonic stage and you are likely harvesting cells from youngsters or adult individuals. It has been quite well demonstrated that adipose derived, umbilical cord and bone marrow MSC after culture in MSC medium for 4-5 passages, should be 90-100% MSC.
https://stemcellres.biomedcentral.com/articles/10.1186/scrt414
Tri-lineage differentiation (cartilage, fat and bone) is a good way to characterize cell type in addition to defining the immunophenotypes.
What is your purpose with these cells?
Good luck
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