28
section. This can be done by using a wedge-tipped sample inlet tube in the flow chamber
(Fulwyler, 1977) or by means of a nozzle which has a cylindrical cross section and a
rectangular orifice (Pinkel, 1982). As discussed above, closed flow chambers having a flow
channel with rectangular cross section may be preferable to nozzles producing a jet in air. It
may be that by choosing the right combination of cross section profiles for the flow channel
and the convergent part of the flow chamber leading into that channel, one may achieve
sufficient orientation of the cells. Further elimination of the orientation artifact will be
obtained by the use of optics which produce a highly convergent beam of excitation light, the
type of optics used in arc lamp-based instruments. A combination of these two approaches,
for example, an arc lamp-based instrument having a flow chamber which orient non-spherical
cell in the flow, may solve this problem.
Organisms larger than 10 - 20 Jlm may sediment so rapidly that it affects the cell density of
the sample and produce some degree of selection of small cells versus large ones. The
swimming of some cells may conceivably cause similar artifacts, for example with phototropic
species. Hence, it may be necessary to stir the sample continuously while it is being injected
into the flow chamber. This can be done by means of a glass coated iron rod in the sample
tube or sample syringe, which is moved by an exterior magnet.
Large cells in particular are exposed to considerable mechanical stress as they pass through
the fluidic system. Narrow tubing with diameters approaching that of the cell produce shear
forces which may tear the cell apart. This effect may be alleviated by reducing the length of
the sample tube and by increasing the bore of this tubing as much as possible without creating
an unacceptably large dead volume. The tubing should preferably be made from a hydrophobic
material, such as Teflon, or its inside should be coated with a hydrophobic material.
Cell disruption may also be caused by accelerations produced in the flow chamber or nozzle.
Thus, the velocity gradients which result when the flow converges into the channel of the flow
chamber or the orifice of the nozzle may constitute a considerable tear on the cells. The flow
chamber or nozzle should be designed to minimize this effect by having a cross section which
decreases gradually over a relatively long distance in such a manner that the velocity gradient
is nearly constant (Dubelaar et al., 1989).
section. This can be done by using a wedge-tipped sample inlet tube in the flow chamber
(Fulwyler, 1977) or by means of a nozzle which has a cylindrical cross section and a
rectangular orifice (Pinkel, 1982). As discussed above, closed flow chambers having a flow
channel with rectangular cross section may be preferable to nozzles producing a jet in air. It
may be that by choosing the right combination of cross section profiles for the flow channel
and the convergent part of the flow chamber leading into that channel, one may achieve
sufficient orientation of the cells. Further elimination of the orientation artifact will be
obtained by the use of optics which produce a highly convergent beam of excitation light, the
type of optics used in arc lamp-based instruments. A combination of these two approaches,
for example, an arc lamp-based instrument having a flow chamber which orient non-spherical
cell in the flow, may solve this problem.
Organisms larger than 10 - 20 Jlm may sediment so rapidly that it affects the cell density of
the sample and produce some degree of selection of small cells versus large ones. The
swimming of some cells may conceivably cause similar artifacts, for example with phototropic
species. Hence, it may be necessary to stir the sample continuously while it is being injected
into the flow chamber. This can be done by means of a glass coated iron rod in the sample
tube or sample syringe, which is moved by an exterior magnet.
Large cells in particular are exposed to considerable mechanical stress as they pass through
the fluidic system. Narrow tubing with diameters approaching that of the cell produce shear
forces which may tear the cell apart. This effect may be alleviated by reducing the length of
the sample tube and by increasing the bore of this tubing as much as possible without creating
an unacceptably large dead volume. The tubing should preferably be made from a hydrophobic
material, such as Teflon, or its inside should be coated with a hydrophobic material.
Cell disruption may also be caused by accelerations produced in the flow chamber or nozzle.
Thus, the velocity gradients which result when the flow converges into the channel of the flow
chamber or the orifice of the nozzle may constitute a considerable tear on the cells. The flow
chamber or nozzle should be designed to minimize this effect by having a cross section which
decreases gradually over a relatively long distance in such a manner that the velocity gradient
is nearly constant (Dubelaar et al., 1989).