Detection of HBD1 peptide in peripheral blood mononuclear cell subpopulations by intracellular flow cytometry.
Production of human beta-defensin1 (HBD1) in response to LPS in monocytes, myeloid dendritic cells and plasmacytoid dendritic cells (PDC) was examined. Since PDC make up only 0.1-0.5% of the peripheral blood mononuclear cell population, we developed a method to determine HBD1 peptide levels using four-color flow cytometry, which can examine several cell surface or intracellular markers at once. Coupled with intracellular flow cytometry, we determined that PDC and monocytes only made significant amounts of HBD1 when exposed to >50ng/ml LPS for 2h. This response was limited to monocytes when ultrapure LPS was used, and was inhibited in PDC by chloroquine treatment.
Ryan
Lisa K
LK
Department of Oral Biology, New Jersey Dental School, University of Medicine and Dentistry of New Jersey, 185 South Orange Avenue, Newark, NJ 07103, USA. ryanlk@umdnj.edu
Diamond
Gill
G
Amrute
Sheela
S
Feng
Zhimin
Z
Weinberg
Aaron
A
Fitzgerald-Bocarsly
Patricia
P
D000906:Antibodies / Q000276:immunology; D000918:Antibody Specificity; D002738:Chloroquine / Q000494:pharmacology; D003713:Dendritic Cells / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D008070:Lipopolysaccharides / Q000494:pharmacology; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism*; D010954:Plasmacytoma / Q000378:metabolism; D011550:Pseudomonas aeruginosa; D013997:Time Factors; D017186:Titrimetry; D023083:beta-Defensins / Q000096:biosynthesis / Q000097:blood* / Q000276:immunology
D000906:Antibodies; C406407:DEFB1 protein, human; D007106:Immune Sera; D008070:Lipopolysaccharides; D023083:beta-Defensins; D002738:Chloroquine