Improvements to parallel plate flow chambers to reduce reagent and cellular requirements

Abstract

Background: The parallel plate flow chamber has become a mainstay for examination of leukocytes under physiologic flow conditions. Several design modifications have occurred over the years, yet a comparison of these different designs has not been performed. In addition, the reagent requirements of many designs prohibit the study of rare leukocyte populations and require large amounts of reagents.

Results: In this study, we evaluate modifications to a newer parallel plate flow chamber design in comparison to the original parallel plate flow chamber described by Lawrence et al. We show that modifications in the chamber size, internal tubing diameters, injection valves, and a recirculation design may dramatically reduce the cellular and reagent requirements without altering measurements.

Conclusions: These modifications are simple and easily implemented so that study of rare leukocyte subsets using scarce or expensive reagents can occur.

Figure 1

Schematic of parallel plate flow chamber configurations. (A) The parallel plate flow chamber as described by Lawrence et al [Reference 7]. (B) The parallel plate available from Glycotech, (C) Schematic of the single pass chamber with value in place for injection of cells or soluble reagents, (D) Schematic for recirculating system with parallel plate flow chamber.

Figure 2

Comparison of measurements in the 3 parallel plate flow chamber configurations. (A) Shear curves generated by perfusing neutrophils over activated with LPS (squares) or unactivated (triangles) EC using the classic (closed triangle and square) or single pass (open triangle or square) Glycotech chambers. The experiment was performed as described in the material and methods section. (B) Constant shear experiments comparing the three different chamber configurations as described in material and methods. Neutrophils were perfused over activated endothelium in either the classic (B1), single pass Glycotech (B2), or recirculation chambers (B3). The number of rolling (black bars) and arrested (clear bars) cells were determined each minute. S.D. is indicated by bars. S.D. bars directed up represent S.D. for rolling cells, and S.D. bars directed down represent S.D. for firm cells; n = 3.