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If you have a UV laser, you may want to check out the following reference for
the identification and isolation murine stem cells. This is a novell approach
which does not rely solely on lineage markers.
RECORD NO.: 96261679
AUTHOR: Goodell MA; Brose K; Paradis G; Conner AS; Mulligan RC
ADDRESS: Whitehead Institute for Biomedical Research, Massachusetts
Institute of Technology, Cambridge 02142, USA.
TITLE: Isolation and functional properties of murine hematopoietic
stem cells that are replicating in vivo.
SOURCE: J Exp Med (I2V), 1996 Apr 1; 183 (4): 1797-806
LANGUAGE: English
COUNTRY PUB.: UNITED STATES
ANNOUNCEMENT: 9610
PUB. TYPE: JOURNAL ARTICLE
ABSTRACT: Hematopoietic stem cells (HSC) are multipotent cells that
reside in the bone marrow and replenish all adult
hematopoietic lineages throughout the lifetime of the
animal. While experimenting with staining of murine bone
marrow cells with the vital dye, Hoechst 33342, we
discovered that display of Hoechst fluorescence
simultaneously at two emission wavelengths revealed a small
and distinct subset of whole bone marrow cells that had
phenotypic markers of multipotential HSC. These cells were
shown in competitive repopulation experiments to contain the
vast majority of HSC activity from murine bone marrow and to
be enriched at least 1,000-fold for in vivo reconstitution
activity. Further, these Hoechst-stained side population
(SP) cells were shown to protect recipients from lethal
irradiation at low cell doses, and to contribute to both
lymphoid and myeloid lineages. The formation of the Hoechst
SP profile was blocked when staining was performed in the
presence of verapamil, indicating that the distinctly low
staining pattern of the SP cells is due to a multidrug
resistance protein (mdr) or mdr-like mediated efflux of the
dye from HSC. The ability to block the Hoechst efflux
activity also allowed us to use Hoechst to determine the DNA
content of the SP cells. Between 1 and 3% of the HSC were
shown to be in S-G2M. This also enabled the purification of
the G0-G1 and S-G2M HSC had a reconstitution capacity
equivalent to quiescent stem cells. These findings have
implications for models of hematopoietic cell development
and for the development of genetic therapies for diseases
involving hematopoietic cells.
MESH HEADINGS: Bone Marrow--growth & development (GD)/cytology (*CY);
Hematopoietic Stem Cells--classification (CL)/cytology
(*CY); Antigens, Ly; Benzimidazoles; Bone Marrow
Transplantation; Cell Division; Cell Separation; Fluorescent
Dyes; Membrane Proteins; Mice; Mice, Inbred C57BL; Radiation
Protection; Spleen--cytology (CY); Staining; Verapamil--
pharmacology (PD); Animal; Female; Support, Non-U.S. Gov't;
Support, U.S. Gov't, P.H.S.
CHEMICAL SUBS: 0 (Antigens, Ly); 0 (Benzimidazoles); 0 (Fluorescent Dyes);
0 (Membrane Proteins); 0 (Sca-1 antigen); 23491-52-3 (HOE
33342); 52-53-9 (Verapamil)
GRANT NO.: HL37569; HL; NHLBI
STANDARD NO.: 0022-1007
DATES: Entered 960805
Best of luck,
Glenn Paradis
MIT Flow Cytometry Core Facility
E17-121A
40 Ames Street
Cambridge, MA 02139
Voice: (617) 253-6454
Fax: (617) 253-3714
Email: gap@mit.edu
> Date: Wed, 15 Jan 1997 14:55:41 -0500
> X-Sender: amixon@pop.nci.nih.gov
> X-Mailer: Windows Eudora Version 2.0.3
> Mime-Version: 1.0
> Content-Type: text/plain; charset="us-ascii"
> To: Cytometry Mailing List <cytometry@flowcyt.cyto.purdue.edu>
> From: Arnold_Mixon@nih.gov (Arnold Mixon)
> Subject: murine stem cells
>
>
> An investigator read somewhere(?) of a panel of monoclonal Abs used
> to identify murine progenitor/stem cells. Of interest is an antibody called
> "lin-neg." Anyone out there heard of it before/know of it's source? It
> sounds like it might be a cocktail of different Abs.
> Thanks,in advanced, to all responses.
>
> Arnold
> Arnold Mixon
> National Institutes of Health
> N.C.I., Surgery Branch
> Flow Cytometry Lab
> Tel.# 301-496-9816
> E-mail: Arnold_Mixon@nih.gov
>
> "Just the FACS Ma'am - I only want the FACS !!"
>
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CD-ROM Vol 3 was produced by Monica M. Shively and other staff at the
Purdue University Cytometry Laboratories and distributed free of charge
as an educational service to the cytometry community.
If you have any comments please direct them to
Dr. J. Paul Robinson, Professor & Director,
PUCL, Purdue University, West Lafayette, IN 47907.
Phone: (765)-494-0757;
FAX(765) 494-0517;
Web