A simple quantitative in vitro macrophage migration assay

An in vitro macrophage chemotaxis model using mouse peritoneal non-elicited resident macrophage cells and chemotaxins containing mediators of non-specific elicitors such as oyster glycogen or sodium caseinate has been described. Macrophage cells accumulation in mouse peritoneal cavity was maximum at 48 hr after injecting (i.p.) oyster glycogen (2.5%) or sodium caseinate (12%), 0.5 ml/mouse. Chemotaxins containing mediators were prepared from these mice by peritoneal lavage and termed as routine 'diluted' cocktail and 'concentrated (3 times)' cocktail. Chemotaxis assays were carried out in a modified Boyden chamber using a 48-well microchemotaxis assembly. In vitro results showed higher macrophage chemotaxis response against the 'concentrated' cocktails as compared to routine 'diluted' cocktail. Macrophages exhibited cell density dependent increase in the responsiveness to chemoattractant and macrophage cell density of 4 x 10(6) per ml concentration in the upperwell was found to be optimum. Macrophage responsiveness was seen better with sodium caseinate cocktail as compared to oyster glycogen in vitro as well as in vivo. DMSO (Dimethyl Sulphoxide) solvent (0.25% conc.) did not interfere with normal macrophage chemotaxis. Both CO2 incubator (5% CO2 in air) and BOD incubator with humidified chamber favoured chemotaxis. In vitro test system described can be used as a model to study the effect of anti-inflammatory compounds directly on the macrophage chemotaxis.