Studies on human peripheral monocyte chemotaxis Part I. A technical study

The human monocyte is an important member of the host defence system, but has been little studied in vitro. In this report, I describe a reproducible and simple method for monocyte chemotaxis in man, using a Boyden chamber. Monocyte suspension was prepared by Ficoll-Hypaque gradient sedimentation. The purity of the suspension was checked with Peroxidase-Giemsa staining. Chemotaxis assay was performed using a 5 μm pore sized Millipore filter(thickness:150 μm) and a Nuclepore filter (thickness:13 μm). When the Millipore filter was used, migrated cells stayed in the filter, while it appeared that cells detached from the filter when the Nuclepore filter was used. In the modified Boyden method using a Millipore filter, more than 3×10(5) monocytes had to be added to the upper room of the chamber, and the duration of 90 minutes was adequate for incubation. Zymosan activated human serum diluted 20 percent was used as the chemoattractant. To estimate chemotaxis in the Millipore filter, migrated cells in the filter were counted at 30 μm depth