Three-dimensional analysis of CD34 sialomucin distribution on cord blood and bone marrow.
Determining the cellular distribution of key adhesion molecules may aid in understanding haematopoietic progenitor/stem cell (HPSC) homing to bone marrow (BM). CD34, a well-characterized marker for blast-like HPSC, is widely used for the isolation and enumeration of HPSC. Functional studies have yet to identify a ligand for CD34. However, growing evidence suggests that CD34 may aid the regulation of HPSC differentiation and modulate the expression of other HPSC adhesion molecules necessary for homing. This study aimed to determine CD34 distribution on umbilical cord blood (CB) and BM. CD34-selected cells were adhered to positively charged gold slides at room temperature, before indirect fluorescent antibody labelling with fluorescein isothiocyanate. Fluorescent distribution was determined by 1-microm interval, confocal laser scanning microscope Z-sections. Initial analysis showed CD34 distributed within peripheral halos and dense pocket regions. The development of three-dimensional imaging software enabled spatial visualization of CD34 distribution on CB and BM in association with differential interference contrast cell image. This showed that CD34 was distributed within peripheral halos, with magnupodia-associated 'meridian-shaped crescents', extending from points of cell-slide adhesion towards the top of the cell. True CD34 distribution, not previously discernible by confocal laser scanning microscopy alone, suggests a possible role in cell adhesion/homing revealed by three-dimensional imaging.
Whiting
Karen A
KA
King-George Laboratory, St George's Hospital Medical School and Kingston University, London, UK.
McGuckin
Colin P
CP
Wertheim
David
D
Pearce
Daniel
D
Pettengell
Ruth
R
D018952:Antigens, CD34 / Q000032:analysis*; D015415:Biomarkers / Q000032:analysis; D001854:Bone Marrow Cells / Q000737:chemistry*; D005312:Fetal Blood / Q000737:chemistry*; D016650:Fluorescein-5-isothiocyanate; D019084:Fluorescent Antibody Technique, Indirect; D005456:Fluorescent Dyes; D006412:Hematopoietic Stem Cells / Q000276:immunology; D006801:Humans; D021621:Imaging, Three-Dimensional; D018613:Microscopy, Confocal
D018952:Antigens, CD34; D015415:Biomarkers; D005456:Fluorescent Dyes; D016650:Fluorescein-5-isothiocyanate