Assessment of total immunoreactive lactoferrin in hematopoietic cells using flow cytometry.
Cell-associated lactoferrin (Lf) was analyzed using a new method involving cell permeabilization, indirect immunofluorescence staining, and flow cytometry. Statistical techniques to evaluate the results for percentage of positive cells, relative fluorescence and homogeneity of Lf distribution were also devised. Most normal adult neutrophils (97.1 +/- 0.3% (SEM), range 92.7-99.6%, n = 41) had brilliant fluorescence homogeneously distributed among the cells. There was significantly greater homogeneity of neutrophil Lf distribution in post-menopausal than pre-menopausal females. In chronic myelogenous leukemia (n = 13) and cord blood (n = 7), fractions of Lf-positive neutrophils were decreased (77.3 +/- 7.5%, range 13.3-96.3%; 71.4 +/- 9.3, range 32.0-95.6%, respectively). Normal monocyte-rich isolates had moderate fluorescence (28.7 +/- 3.6%, range 9.3-76.8%, n = 22). Among blood lymphocyte-rich preparations, 13.1 +/- 1.3% of cells had weak positivity (range 4.9-26.6%, n = 19); monoclonal B and T lymphocytes had similar parameters. No other cells had detectable Lf. Our results were significantly correlated with those obtained manually (r = 0.98, P less than 0.001), and are consistent with Lf quantity and distribution determined using other methods.
Butler
T W
TW
Veterans Administration Medical Center, Birmingham, AL 35233.
Heck
L W
LW
Huster
W J
WJ
Grossi
C E
CE
Barton
J C
JC
D000328:Adult; D000818:Animals; D000911:Antibodies, Monoclonal; D000937:Antigen-Antibody Reactions; D001773:Blood Cells / Q000032:analysis*; D002448:Cell Adhesion; D002469:Cell Separation / Q000379:methods; D005260:Female; D005312:Fetal Blood / Q000032:analysis; D005434:Flow Cytometry / Q000379:methods*; D005455:Fluorescent Antibody Technique; D006801:Humans; D007231:Infant, Newborn; D007781:Lactoferrin / Q000097:blood*; D007782:Lactoglobulins / Q000097:blood*; D007963:Leukocytes, Mononuclear / Q000032:analysis; D008297:Male; D051379:Mice; D009504:Neutrophils / Q000032:analysis
D000911:Antibodies, Monoclonal; D007782:Lactoglobulins; D007781:Lactoferrin