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Techniques

Dead volume

Dead volume should be minimized in order to switch solu-

tions as rapidly as possible. Dead volume for this purpose refers to

the volume in the single piece of tubing be tween the manifold and

fi nal destination (perfusion chamber, 96-well plate, etc.) A certain

amount of time must pass for a new liquid to clear the previous one

from this fi nal piece of tubing. Solution switching takes place at the

manifold - assuming each channel's tubing has been primed from

its valve to this point. Therefore, the manifold should be positioned

as close to the ultimate destination as possible, while the valves can

be some distance away. The micro-manifolds available from Auto-

Mate Scientifi c are designed to be inserted without tubing directly

into orifi ces in some perfusion chambers. This absolutely minimizes

dead volume – resulting in switching times of under a second.

AutoMate Perfusion Systems include a blue disposable fl ow

regulator which will increase dead volume if used improperly. The

regulator can either be eliminated if controlling fl ow rate is not

necessary, or positioned before the manifold to eliminate dead vol-

ume. However, this second method only controls the fl ow rate of

individual solutions (instead of all solutions when located after the

manifold). Therefore, the researcher can either limit the fl ow of a

single liquid, or purchase fl ow regulators for each line.

A fi nal method for reducing dead volume is to use smaller

bore tubing, although this may also reduce fl ow rate re sult ing in the

same switching time as for larger tubing.

Small bore, Tefl on tubing

AutoMate Scientifi c's micro-manifolds use 1/16" o.d. tube

ports which can either be used with this size tubing, or enlarged up

to 1/16" i.d for valve hose barb fi ttings. Please inquire about Au-

to Mate's Tefl on tubing and nut and ferrule valve fi ttings for small

bore use. Also see the micro-manifold instructions on page 7.

Cutting Tefl on tubing at an angle makes it easier to insert.

Backfl ow

When a manifold is used in a plumbing arrangement, none of

its ports can be left unconnected, or liquid will simply fl ow back-

wards out of the opening. Small, green hose clamps are in clud ed

with AutoMate Perfusion System drippers which can be easily at-

tached to short pieces of tubing to act as plugs for un used holes.

Likewise, one must be careful not to open any valve whose

reservoir is completely empty or infl ow tube is disconnected. Or-

di nary so lu tion pressure in other lines will force liquid back up the

outfl ow tubing, through the valve, and either back into the reservoir

or out the dis con nect ed infl ow onto the fl oor. Again, clamp-off any

un used lines at the manifold or be sure all valves are closed while

changing con nec tions.

Syringe reservoirs

AutoMate offers 30ml syringes (photo on page 3) as al ter na -

tives to its 100ml polypropylene reservoir cups. Connected directly

into luer lock fi ttings in the valves, this method eliminates wasted

solution in tubing between the reservoirs and valves. Small 2-way

stopcocks are included when this option is ordered. These are

placed between the syringe and valve fi tting in case syringes must

be disconnected before completely empty. Simply loosen the luer

lock from the valve fi tting, and remove the syringe and stopcock to-

gether. These stop cocks are also useful if there is a chance of a valve

opening after its reservoir is empty (see above paragraph).

AutoPrime

System

™

The AutoPrime Perfusion System is de-

signed to deliver oxygenated (or other gas-sat-

u rat ed) solutions without ad min is ter ing 'stale'

liquid which has remained sta tion ary in the

tubing long enough to lose its ox y gen ation.

Unless one uses 'hard-walled' (i.e. Tef-

lon) tubing, gas will escape through the

tubing leaving the stale solution with a

ruined pH and gas con cen tra tion. The 8-

channel AutoPrime System uses eight valves

per four liquids, with the second set used as

waste