U.S. patent number 3,764,041 [Application Number 05/083,967] was granted by the patent office on 1973-10-09 for microdispensing process and apparatus.
This patent grant is currently assigned to G. D. Searle & Co.. Invention is credited to Hans Noll.
United States Patent |
3,764,041 |
Noll |
October 9, 1973 |
MICRODISPENSING PROCESS AND APPARATUS
Abstract
A process and apparatus for the precise dispensing of very small
volumes of liquids or gels wherein the means delivering such
liquids or gels to collection vessels is rinsed at the end of each
dispensing interval, thereby cutting a continuous stream of such
materials into fractions of equal volume regardless of drop size. A
process and apparatus which permits addition of reagent fluids to
each of the collection vessels in conjunction with the dispensing
of the fractions to be studied.
Inventors: |
Noll; Hans (Evanston, IL) |
Assignee: |
G. D. Searle & Co. (Skokie,
IL)
|
Family
ID: |
22181815 |
Appl.
No.: |
05/083,967 |
Filed: |
October 26, 1970 |
Current U.S.
Class: |
222/1; 222/132;
73/864.22; 222/148; 422/401 |
Current CPC
Class: |
G01N
35/1016 (20130101) |
Current International
Class: |
G01N
1/00 (20060101); G01n 001/00 () |
Field of
Search: |
;222/1,145,132,148,149,193,420,421,488 ;239/112,424 ;141/90,91,105
;73/425.4 ;23/259 |
References Cited
[Referenced By]
U.S. Patent Documents
Primary Examiner: Reeves; Robert B.
Assistant Examiner: Scherbel; David A.
Claims
I claim:
1. Apparatus for dispensing precise volumes of a fluid which is a
member of the group consisting of a liquid and a gel
comprising:
a dispensing conduit connectable to a reservoir of said fluid and
terminating in a dispensing tip;
means adjacent to at least a segment of the dispensing conduit,
providing a passageway, and communicating with a rinsing liquid
supply,
dispensing pump means communicating with said dispensing conduit
for delivering a predetermined volume of said fluid dropwise
through said dispensing tip, and
discontinuous flushing pump means communicating with said
passageway for delivering a rinsing liquid at said dispensing tip
and adapted to be actuated when delivery of said predetermined
volume is substantially complete so as to flush from said
dispensing tip fluid collected thereon; whereby the rinsing liquid
flushes from the dispensing tip any fluid collected externally
thereon to reproducibly complete the volume of fluid dispensed.
2. Apparatus according to claim 1 in which said conduit comprises a
hollow needle and said first-mentioned means comprises a
cylindrical tubular casing having an inner diameter greater than
the outer diameter of said needle,
said needle being positioned within said casing so that at least
the dispensing tip extends outwardly from the bottom thereof.
3. Apparatus according to claim 2 in which the flushing pump means
delivers the rinsing liquid under pressure and the difference
between the inner diameter of the casing and outer diameter of the
needle is such as to be effective to prevent flow of the flushing
liquid between flushing cycles.
4. Apparatus according to claim 3 in which said flushing pump means
is provided with a hollow side arm projecting from the casing and
communicating with said passageway at a point adjacent to a top
portion of said passageway.
5. Apparatus according to claim 4 and comprising further a cap
member sealing the top portion of said passageway and accommodating
said needle therethrough to maintain the same coaxially within the
casing.
6. Apparatus according to claim 2 and comprising further reagent
dispensing means for dispensing a reagent during the dispensing
cycle of the liquid or gel responsive to a pre-determined
signal.
7. A process for obtaining precise test fractions from a reservoir
containing a fluid which is a member of the group consisting of a
liquid and a gel characterized by the steps of:
dispensing dropwise a predetermined volume of the fluid for a
predetermined cycle through a small-diameter conduit having a
dispensing tip and into a collecting vessel, and
intermittently delivering at said dispensing tip when the
dispensing of said predetermined volume is substantially complete a
rinsing liquid flushing from the dispensing tip and into the
collecting vessel any fluid collected on said dispensing tip and
thereby reproducibly completing each collecting cycle.
8. The process of claim 7 wherein the rinsing liquid comprises a
reagent for use in chemical analysis of the test fraction.
9. The process of claim 7 further characterized by the step of
dispensing into the test fraction a liquid scintillation reagent
simultaneously with the delivery of the rinsing liquid.
10. In an apparatus for continuously and automatically obtaining
precise test fractions from a liquid or gel reservoir for further
analysis and including a rack containing a plurality of collection
vessels, a delivery head mounted above said vessels and means for
moving delivery of liquid or gel from one vessel to the next at a
pre-determined dispensing cycle, the improvement comprising:
a supporting block attached to said head;
a small-diameter conduit having a dispensing tip;
a tubular casing of larger inner diameter sheathing a portion of
said conduit and providing an annular passageway therealong;
means connecting the conduit to the casing in coaxial relationship
so that the dispensing tip extends beyond the bottom of the
casing;
a side arm projecting from the casing and communicating with said
passageway;
means in said block cooperating with said conduit, casing and side
arm to support the same in readily removable relationship;
means connecting said side arm to a flushing liquid source;
pump means for delivering flushing liquid to the side arm; and
timer means for automatically actuating the pump means at a
pre-determined time in each dispensing cycle,
whereupon the flushing liquid flows through said passageway and
rinses from the dispensing tip any liquid or gel collected thereon
to complete each test fraction.
11. Apparatus according to claim 10 in which the difference between
the inner diameter of the casing and the outer diameter of the
conduit is such as to be effective to prevent flow of the flushing
liquid between flushing cycles.
12. Apparatus according to claim 11 in which the flushing liquid
comprises a reagent for use in chemical analysis of the test
fractions.
13. Apparatus according to claim 10 and comprising further a
reagent spout;
means for removably and adjustably connecting said reagent spout to
the supporting block;
means connecting the reagent spout to a liquid scintillation
reagent source; and
second pump means for delivering the scintillation reagent to said
spout;
said timer means automatically actuating said second pump means to
dispense into the test fraction simultaneously with the delivery of
the flushing liquid a fixed amount of the liquid scintillation
reagent for use in radioactive assay of the test fractions.
Description
This invention relates to precision dispensing of liquids or gels
and more particularly to improved apparatus and methods for use in
laboratory fraction collecting and testing.
BACKGROUND OF THE INVENTION
An important operation in biomedical research and pharmaceutical
manufacturing is the dispensing of identical volumes of liquids or
gels into collection vessels or collectors by a process called
fraction collection. In a typical operation of this type, small
volumes of liquids or gels are dispensed mechanically and
continuously into a series of collection vessels or tubes. The
smallest quantity of liquid or gel that can be delivered by
existing devices is determined by the size of the drop forming at
the tip of the delivery tube or needle above the collection vessel.
As a result, the obtaining of a precise fraction and the
reproducibility thereof is frequently impossible.
Current fraction collectors designed for use in combination with
chromatography, electrophoresis or ultracentrifugation procedures
cut fractions by drop counting, by constant volume metering devices
operated by mechanical siphoning or automatic valve action, or by
interval timing. All of these methods, in which the liquid is
allowed to drop into a series of tubes by gravity, are
unsatisfactory for the collection of small equal fractions (0.5 ml
and less), because (1) the desired fraction size does not
correspond to a whole number of drops, (2) drops are lost between
tubes at the moment of changing fractions, (3) the drop size
changes with the concentration of solutes in the liquid, and (4) a
relatively large and variable portion of the fraction volume
remains at the tip of the delivery tube or needle. Moreover, the
resolution often is determined by the smallness of the fractions
that can be collected with precision. This applies to all
procedures in which the detection and quantitative analysis of the
substances that have been separated into narrow and closely spaced
bands or zones must be carried out by discontinuous sampling. Thus,
the separation into two closely spaced bands will only be
detectable if a sufficient number of small fractions can be
collected for analysis.
Fractionation into smaller volumes can be achieved by pipetting (as
in manual laboratory practice) by touching the tip of the pipette
to the surface of the receiving vessel or tube or by immersing the
tip below the surface of a diluent fluid present in the receiving
vessel. Such a pipette and process is described in United States
Pat. No. 2,991,647. The first method, however, entails obvious
losses and the second causes contamination of subsequent vessels
with the fluid present in the preceding tube. Both methods are
cumbersome and difficult to adapt for automatic operation.
The problem of small volume collection fractioning can, of course,
be alleviated by diluting the solution to be dispensed and thereby
increasing the delivery volume so that it becomes large relative to
drop size. There are, however, many applications in which this is
not feasible, as in the continuous fractionation of inhomogeneous
media (exemplified by density gradients and gels in centrifugation
and electrophoresis) used for the separation and partitioning of
substances.
With existing equipment it has not been possible to process by an
automated operation density gradients of small volume (3-20 ml)
with acceptable resolution and precision. A well-known commercial
fraction collector accommodating liquid scintillation counting
vials permits the automatic fractionation of density gradients of
relatively large volume (over 20 ml), but the usefulness of such
instruments is limited because of new low-volume,
fast-centrifugation techniques and the need for independent
addition of the required scintillation reagents.
DESCRIPTION OF THE INVENTION
My invention comprises a novel configuration of components
providing for the precise dispensing of very small volumes of
liquids or gels, thereby overcoming the limitations of automatic
and manual fraction collecting mentioned above. The apparatus and
process of my invention achieves this precision by removing even
fractional drops from the tip of the tube or needle delivering the
liquid or gel to the collection vessels by flushing this tip with a
small volume of a suitable rinsing liquid added on command from a
supply reservoir, immediately preceding the advance of the delivery
tip to the next vessel. The rinsing liquid may comprise any liquid
physically and chemically compatible with the fraction such as
distilled water or suitable organic solvents. Where subsequent
procedures require the addition of chemical reagents, the rinsing
liquid may comprise or include such reagents.
My invention also comprises a delivery spout which may be operated
so that reagents necessary for the liquid scintillation assay of
collected fractions are dispensed simultaneously with the
fractions, thereby eliminating time-consuming and inefficient
separate addition thereof.
The invention is adaptable for automated or manual collection
systems and, when used in conjunction with a commercial moving
fraction collector, timers, and commercial pumps, makes possible a
completely automatic system capable of processing a liquid or gel
for radioassay or chemical testing. Substantial economies of time
are thus achieved as well as greater accuracy and improved
radioassay resolution, since resolution increases with the smaller
size fractions obtainable by use of my invention.
Typical applications of the automatic microdispensing fractionation
and processing system made possible by my invention are the
processing of liquid density gradients and acrylamide
electrophoresis gels for radioassay and the automatic chemical
testing of fractions obtained from chromatographic columns, density
gradients or acrylamide gels. The fractionation of gels by existing
techniques is similar to the collection of liquids and subject to
the same limitations. By applying the discontinuous flushing action
and the reagent addition of my apparatus, the same increase in
accuracy and savings in time is obtainable for the processing of
gels.
Accordingly, it is an object of my invention to provide a process
and apparatus for the precise dispensing of very small volumes of
liquids or gels.
It is a further object of my invention to provide a process and
apparatus of the character described and having means for the
automatic and simultaneous addition of reagents which may be
required for subsequent radioassay or chemical testing of samples
collected during fractionation.
It is another object of my invention to provide an apparatus
containing a liquid or gel microdispensing device in combination
with a reagent dispensing device, such apparatus being adaptable to
fit any existing automatic fraction collector.
It is a still further object of my invention to provide an
apparatus and process combining microdispensing of liquids and gels
with dispensing of reagents which substantially reduces the time
required for fraction collecting and processing.
These and other important objects of the invention will become
apparent from the following description taken in conjunction with
the drawings illustrating a preferred embodiment wherein:
FIG. 1 is a front elevation view of a liquid or gel microdispensing
and reagent dispensing apparatus embodying the invention, and
FIG. 2 is an enlarged vertical cross section view of the dispensing
assembly of my microdispensing apparatus.
Referring specifically to FIG. 1, my liquid or gel microdispensing
reagent dispensing apparatus comprises an L-shaped retainer block
10, that attaches by means of a baseplate 70 to the lower portion
of any known liquid or gel delivery head (not shown). The baseplate
70 has suitable slits for accommodating set screws 71 to enable
adjustments of the apparatus in the direction of travel and
laterally with respect to the center of the collection vessels, as
required. The retainer block 10 has a vertical bore 12 whose upper
portion is open-sided to afford a vertical channel 14 and an
open-topped channel 16 communicating therewith to form an open
acute-angle joint. The bore and two channels cooperate to removably
support therein the liquid dispensing assembly 18.
Dispensing assembly 18 comprises a hollow needle or small-diameter
conduit 20 which is connectable at its upper end, as by suitable
tubing 22, to a liquid or gel supply and which terminates at its
bottom in a dispensing tip 24. A cylindrical tube or casing 30
having an internal diameter greater than the external diameter of
the needle 20 is positioned in sheath-like manner over the needle
20 to provide an annular passageway 32 therearound. Passageway 32
is closed at its upper end by a removable cap member 34 having a
central aperture 36 which supports the needle 20 coaxially within
the passageway 32. The open bottom end 38 of the casing 30 is
positioned above the dispensing tip 24 for reasons which will
subsequently become apparent.
A tubular side arm 40 projects at an acute angle from the casing 30
and communicates with the passageway 32, said side arm being
connectable, as by suitable tubing 42, with a rinsing liquid supply
and flushing pump. It should thus be noted that the entire assembly
18 is supported by the contour-accommodating bore 12 and channels
14 and 16 so that the same may be readily removed for purposes of
cleaning, maintenance and replacement.
The foot of the L-shaped block is formed with a vertical channel or
cut-out defined by a vertical wall 44 and opening to the facing
edge 45 of the foot. This channel is adapted to slideably and
removably receive an inset block 46 which carries, through a
vertical hole 48 provided therein, a curved delivery spout 50
having a vertical section 52. Delivery spout 50 is connectable, as
by suitable tubing 54, to a scintillation liquid reagent source and
a pump. Lateral positioning of the delivery spout 50 is adjustable
by means of a set screw 56 adapted to track in a horizontal groove
58 formed in a side wall of inset block 46, while vertical and
radial positioning of the spout is adjustable by means of a set
screw 60 carried by the inset block and adapted to bear against
vertical section 52 of the spout. The spout thus may be positioned
precisely with relation to dispensing tip 24 and the receiving
vessels as dictated by the particular conditions obtaining.
In the operation of my microdispensing and reagent dispensing
process, the drops of liquids to be dispensed and collected form at
the tip 24 of the needle 20 above the collecting vessels. At the
pre-set interval in each dispensing cycle, the flushing pump is
activated to pump a charge of rinsing liquid through the side arm
to and into the passageway 32 where it flows downwardly and
circulates around the needle 20 to emerge at the casing bottom 38.
Upon emergence, the rinsing liquid surrounds and flows down the
protruding dispensing tip 24 of the needle 20 in a stream, flushing
from the tip the drop or fractional drop formed thereat. The
relative diameters of the needle 20 and passageway 32 are in some
measure a function of the surface tension of the rinsing liquids
and are determined to prevent unwanted flow of the rinsing liquids
between flushing actions.
The invention makes possible the development of a totally automated
system for use in any of the analytical procedures described
hereinabove. In such a system, the microdispensing and reagent
dispensing apparatus is combined with a commercial fraction
collector, which moves the collecting vessels relative to a
stationary delivery system or which moves the delivery head over a
rack of collecting vessels. The microdispensing apparatus will be
interposed between the delivery system and the collecting vessels.
Also included are such known elements as a reagent supply and a
reagent pump, a rinsing liquid supply and pump, a first timing
means regulating the collecting interval, and a second timing means
regulating the delivery of the rinsing liquid and the scintillation
reagent, when required. It should of course be appreciated that
where the particular procedure involves chemical tests of the
fractions, the rinsing liquid may contain, or comprise solely, the
reagent or reagents required.
For proper operation of the automated system described, the
flushing and reagent pumps are of the one-shot repetitive-action
type. The action of the pumps is synchronized with the advancement
of the delivery head or collecting vessels depending on the
fraction collector used in such a way that the pumping action is
triggered and the liquid addition completed immediately before
advancement to the next collection vessel. This has been
accomplished by using the signal of the first timing means, such as
an interval timer normally controlling the advancement to the next
collecting vessel in the standard fraction collecting unit, and
feeding the signal into a secondary timing means, such as a delay
or slave timer, which in turn actuates the pumps. The slave timer
is set for an interval that is a few seconds shorter than the
collection interval. Hence, with each advancement to the next
collecting vessel, the slave timer is started. At the end of the
interval, or a few seconds before the next advancement, the slave
timer actuates the pumps and the rinsing liquid and/or reagent are
introduced into the microdispensing and reagent dispensing
apparatus.
The retainer block and inset block preferably are constructed of a
rigid, lightweight, easily shaped or cut material such as metal,
plastic or wood in order to properly support and maintain in
position the various dispensing means. The dispensing assembly 18
may be made of any suitable non-reactive and easily cleanable
material such as stainless steel or glass.
While the invention has been described in relation to a preferred
embodiment thereof, it will be apparent to those skilled in the art
that the structural details are capable of wide variation without
departing from the principles of the invention.
* * * * *