14. C E L L A G G R E G A T E S
495
explanted to a plasma clot, underwent histogenesis. The advantages
here are operational simplicity and an apparent minimal loss of cells.
However, even gentle centrifugation may be harmful to dissociated
cells. It may result in entrapment of the dissociating agents (trypsin,
Versene, etc.) and degradation products between the packed cells, and
may rapidly exclude most ofthe cells from access to the culture medium,
furthermore, centrifugation imposes stratification patterns on the
packed cells which may not be at all related to their manner of association in tissues. Therefore results thus obtained must be carefully evaluated particularly in work on cellular affinities, sorting out, and
preferential localization of cells. Packing of cells by centrifugation
is of limited usefulness in studies on differential adhesion of cells or on
factors that affect the binding of cells into aggregates.
2. Aggregation by Rotation
In aggregation by rotation, the dispersed cells are concentrated by
gently rotating the suspensions in Erlenmeyer flasks on a gyratory
shaker. The spinning flow of the liquid assures a high frequency of
random collisions, without forceful packing, and if the cells are mutually
cohesive they form aggregates ("rotation-mediated aggregation") (Moscona,
1960, 1961a; for aggregation of slime molds in roller tubes see Gerisch,
1960). Initial clusters continue to collect cells and to merge until reaching a size characteristic for the given cell population and conditions,
whereupon no further mergers take place. Instead of the sedimentation
which occurs in self-aggregation, in this system cells and aggregates
remain in suspension all the time; their collisions and contacts are
therefore largely independent of the migratory capacities or growth of
the cells and ofthe factors that may affect these. The chief advantage of
aggregation by rotation is that all pertinent procedural parameters can
be readily controlled and standardized. Consequently, results are consistently reproducible and the system lends itself to quantitative studies
on cell interactions and differentiation under well defined conditions.
Formation of aggregates by rotation reflects the combined action of:
(a) the concentric flow of the medium that tends to sweep cells and
emerging aggregates into the vortex, to increase collision frequency, and
so to promote aggregation; (b) the shearing forces that tend to separate
and disperse the cells and to reduce collision frequency; and (c) the
mutual adhesiveness of the cells under the given conditions, (a) and (b)
are related to speed of rotation in that (a) decreases and (b) increases
at higher speeds; at a given speed of rotation only those cells form
aggregates which, upon contact with each other, become mutually
attached firmly enough to resist the effect of the shearing forces at this
speed. The number and size of resulting aggregates represent therefore
495
explanted to a plasma clot, underwent histogenesis. The advantages
here are operational simplicity and an apparent minimal loss of cells.
However, even gentle centrifugation may be harmful to dissociated
cells. It may result in entrapment of the dissociating agents (trypsin,
Versene, etc.) and degradation products between the packed cells, and
may rapidly exclude most ofthe cells from access to the culture medium,
furthermore, centrifugation imposes stratification patterns on the
packed cells which may not be at all related to their manner of association in tissues. Therefore results thus obtained must be carefully evaluated particularly in work on cellular affinities, sorting out, and
preferential localization of cells. Packing of cells by centrifugation
is of limited usefulness in studies on differential adhesion of cells or on
factors that affect the binding of cells into aggregates.
2. Aggregation by Rotation
In aggregation by rotation, the dispersed cells are concentrated by
gently rotating the suspensions in Erlenmeyer flasks on a gyratory
shaker. The spinning flow of the liquid assures a high frequency of
random collisions, without forceful packing, and if the cells are mutually
cohesive they form aggregates ("rotation-mediated aggregation") (Moscona,
1960, 1961a; for aggregation of slime molds in roller tubes see Gerisch,
1960). Initial clusters continue to collect cells and to merge until reaching a size characteristic for the given cell population and conditions,
whereupon no further mergers take place. Instead of the sedimentation
which occurs in self-aggregation, in this system cells and aggregates
remain in suspension all the time; their collisions and contacts are
therefore largely independent of the migratory capacities or growth of
the cells and ofthe factors that may affect these. The chief advantage of
aggregation by rotation is that all pertinent procedural parameters can
be readily controlled and standardized. Consequently, results are consistently reproducible and the system lends itself to quantitative studies
on cell interactions and differentiation under well defined conditions.
Formation of aggregates by rotation reflects the combined action of:
(a) the concentric flow of the medium that tends to sweep cells and
emerging aggregates into the vortex, to increase collision frequency, and
so to promote aggregation; (b) the shearing forces that tend to separate
and disperse the cells and to reduce collision frequency; and (c) the
mutual adhesiveness of the cells under the given conditions, (a) and (b)
are related to speed of rotation in that (a) decreases and (b) increases
at higher speeds; at a given speed of rotation only those cells form
aggregates which, upon contact with each other, become mutually
attached firmly enough to resist the effect of the shearing forces at this
speed. The number and size of resulting aggregates represent therefore