Note: Descriptions are shown in the official language in which they were submitted.
3~9
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AUTOMATIC PIPETTING APPARATUS
BACKGROUND OF THE INVENTION
Field of the Invention
This invention is related in general to the field of
applylng fluid samples to analysis strips. In particular,
this invention relates to an automatic pipetting apparatus
for applying multiple fluid samples to a microporous
~0 support medium such as a cellulose acetate or agarose
strip ~hich may be used in the field of zone
electrophoresis and with other separation techniques
including the field of thin layer chromatography. Zone
electrophoresis is the science of moving charged particles
~n an electric field through a solid or semi-solid medium.
The technique is most commonly used in medical research
and medical laboratories for analyzing various blood
proteins.
Description of the Prior Art
~n the electrophoresis technique, a blood or other
fluid sample is applied to a support medium ~hich is then
subjected to an electric field so as to separate the
components of the sample. The support media used in the
electrophoresis process includes cellulose acetate, agar,
agarose and acrylamide gels. In laboratory work it is
desirable that a plurality of samples be applied to the
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support medium such that each of the samples may be
subjected to the electric field at the same time.
Tho samples may be applied to the support medium one
at a time in serial fashion ~ith a hand pipetter, but the
hand pipetter must be rinsed ~ith a cleansing agent and
blotted before a ne~ sample is aspirated and then applied
to the strip.
Applicators have been designed to apply fluid samples
simultaneously or in "parallel" to the strips. Such
applicators are described at page 61 of the General
Products Catalog for 1984-19B5 of Helena Laboratories ~ith
offices in Beaumont, Texas. Such applicators may apply
eight, t~elve or more samples to a microporous support
medium and have the advantage of making the
electrophoresis technique easier and more reproducible.
The applicators kno~n prior to this invention ho~ever
have been essentially non-automatic applicators and
required cleaning of the applicator tips after each
application to the support medium.
Automatic dispensing systems are known in the prior
art. For example, a system sold under the trademark "~Tell
Washers" of BioTech Instruments, Inc. of Burlington,
Vermont provides an alignment mechanism by ~hich a ro~ of
eight or t~elve barrels may be positioned above one of a
plurality of ro~s of ~ashing vials or wells. Automation
in the system provides selection of dispensing fill
volumes, soaking times and number of wash cycles.
12~36389
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None of the prior art ho~ever has provided an
apparatus ~or automatically filling a plurality of
pipetters from a respective plurality of fluid chambers
and then precisely applying such fluid samples from each
pipette to a support medium. Another disadvantage of the
prior art systems is that there has been no means for
automatically ~ashing and cleaning the barrels during each
cycle time so as to prevent contaminatio~ of each of the
barrels during application of a ne~ plurality of fluid
samples to a ne~ support medium.
Another disadvantage of the prior art is that there
has been no means for precisely automatically applying a
very small amount - of the order of one micro liter of
sample liquid - to a support medium.
Another disadvantage of the prior art is that there
has been no means for precisely automatically diluting a
very small amount - of the order of one micro liter - of
sample fluid ~ith a diluting liquid, and precisely
applying a very small amount of the diluted sample to a
support medium.
Accordingly the invention seeks to provide an
automatic pipetter apparatus for aspirating from a
plurality of sample chamber wells into a corresponding
plurality of pipetters and then applying such samples
precisely to~ a microporous support medium to be used in
electrophoresis or thin layer chromatography.
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~Z~3ti3~39
Further the invention seeks to provide an
automatic pipetting apparatus ~hich not only aspirates and
dispenses sample fluids onto a support medium strip such
as cellulose acetate or agarose, but also flushes,
cleanses, rinses and blots the tips of the barrels ~ith an
appropriate cleaning fluid before and after each
application of the sample fluid to the support medium.
Still further the inve~tion seeks to provide an
automatic pipetting apparatus by ~hich p~sitive
displacement pipette barrels and plungers are controlled
to precisely apply a very small sample of fluid to a
support medium.
Further still the invention seeks to provide
automatic plpetting apparatus for precisely automatically
diluting a very small amount of sample fluid with a
diluting liquid and precisely applying a very small amount
of the dlluted sample to the support medium.
SUMMARY OF THE INVENTION
-
Accordingly, in one embodiment of
the invention there is provide,d
an automatic pipetting apparatus ~hich generally includes
a base and a sample plate disposed on the base and a
pipette frame including a vertical support for supporting
the frame from the base above the sample plate. The
sample plate includes a ro~ of individual liquid sample
chambers and a lateral application space longitudinally
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separated from the li~uid chamber ro~. The lateral
application space is adapted to receiv~ a microporous
support medium. A mounting plate is carried by the
pipette frame.
The apparatus includes translation means for
effecting relative longitudinal movement of the pipette
frame and sample plate, and vertical translation means for
effecting rela ive vertical movement of the mounting plate
and the sample plate.
A plurality of microsyringe barrels are removably
~ecured to the mounting plate. The barrels are spaced
corresponding to the spacing of the liquid chambers of the
sample plate. A plurality of micro-plungers are provided,
one each movably disposed in one of the barrels. A
plunger translation means is provided for moving the
plunger vertically within the barrels.
Signalling means are provided for generating
longitudinal signals representative of the relative
longitudinal orientation of the pipette frame with
respect to the sample plate, for generating mounting plate
signals representative of the vertical orientation of the
mounting plate relative to the sample plate and for
generating plunger signals representative of the
orientation of the plungers relative to the barrels.
A programmed digital computer is provided responsive
to the longitudinal signals, the mounting plate signals
and to the plunger signals for generating a sequence of
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128G3~39
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control signals to the longitudinal translation means, to
the vertical translation means and to the plunger
translation means to aspirate a predetermined amount ~f
liquid ~rom the sample chambers into the respective
pipette barrels, and to apply the liquid samples in the
barrels onto corresponding spaces or "spots" on the
microporous support medium when placed on the lateral
application space of the sample plate.
~nother embodiment of the invention includes a base
having a track disposed longitudinally on it. A carriage
i8 longitudinally movably disposed on the track and
carries a sample plate ~hich is removably disposed on the
carriage. The carriage plate includes a lateral ro~ of
individual liquid sample chambers and a lateral
application space ~hich is longitudinally separated from
the liqu~d chamber ro~. The 1ateral application space is
adapted to receive a microporous support medium such as a
cellulose acetate or agarose strip used in electrophoresis
or thin layer chromatography.
A pipette assembly is mounted vertically on the base
above the carriage and the sample plate. The pipette
assembly includes vertical mounting posts separated
laterally from each other and secured to the base. A
mounting plate assembly is slidably guided by the posts
and is disposed laterally ~ith respect to the sample
plate.
.. . . . .
3~3
The mounting plate assembly includes a mounting plate
having slidable guides disposed about the posts. A
pipette bar is fixed to the mounting plate. A plurality
of microsyringe barrels are provided in a ro~ on the
pipette bar ~ith their heads secured thereto~ The
microsyringe barrels are spaced corresponding to the
spacing of the liquid chambers on the plate. The barrels
are hollo~, each barrel having a lower tip.
A plunger bar is vertically movably disposed above
the tip bar and has a plurality of micro-plungers secured
thereto. Each of the micro-plungers are movably disposed
~ithin a corresponding barrel of the microsyringes. A
plunger actuator plate carried by the mounting plate is
vertically movable ~ith respect to the mounting plate.
The actuator plate is removably secured to the plunger
bar.
Translation and signalling means are provided for
moving the carriage longitudinally for~ard and back~ard
beneath the mounting plate assembly and generating
carriage position ~ignals indicative of the carriage
position. A translation and signalling means is provided
for moving the mounting plate assembly up and do~n ~ith
respect to the base and generating mounting plate position
signals indicative of the mounting plate position. A
translation and signalling means for moving the plunger
bar up and do~n ~ith respect to the mounting plate and
lZ~j3fi~
generating plunger bar position signals indicative of the
plunger bar position is provided.
A programmed microcomputer is provided responsive to the
carriage position slgnals, to the mounting plate position
signals, and to the plunger bar position signals for generating
a sequence of control signals to the translation means for
moving the carriage, the translation means for moving the
mounting plate, and the translation means for moving the
plunger bar so as to aspirate a first predetermined amount of
liquid from the sample chambers into the respective pipette
barrels and then to apply the liquid in each of the pipette
barrels onto corresponding spaces of the support medium when
placed on the lateral application space of the sample plate.
Another aspect of the invention provides a sample plate
adapted for use with automatic pipetting apparatus comprising a
lateral row of individual liquid sample chambers and a lateral
application space longitudinally separated from the liquid
chamber row. The lateral application space is adapted to
receive a microporous support medium and includes a raised
portion and a lower portion, the row of individual liquid
sample chambers being disposed on the raised portion, and the
lateral application space disposed on the lower portion
and further comprising a wash well and a waste well
longitudinally spaced from each other and from the sample
chambers.
A further aspect of the invention comprehends a sample
plate adapted for use with automatic pipetting apparatus
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~Z~36389
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comprising a lateral row of individual liquid sample chambers
and a lateral application space longitudinally separated from
the liquid chamber row, the lateral application space being
adapted to receive a microporous support medium, and further
comprising a wash well and a waste well longitudinally spaced
from each other and from sample chambers. There is further a
longitudinal blotting space for applying a lateral blotting
paper strip, the blotting space longitudinally separated from
the sample chamber row, the wash well, the waste well and the
lateral application space.
Still another aspect of the invention pertains to
apparatus adapted for use with automatic pipetting of liquid
samples to be subjected to electrophoresis comprising a sample
plate including a lateral row of individual liquid sample
chambers and a lateral application space longitudinally
separated from the liquid chamber row, the lateral application
space having disposed thereon a removable microporous support
medium adapted for electrophoretic analysis after a liquid
sample has been deposited thereon. The sample plate includes
an upper portion and a lower portion, the row of individual
liquid sample chambers being disposed on the raised portion
and the lateral application space being disposed on the lower
portion.
BRIEF DESCRIPTION OF THE DRAWINGS
The aspects, advantages and features of the invention will
become more apparent by reference to the drawings which are
appended hereto and wherein like numerals indicate like parts
and wherein an illustrative embodiment of the invention is
shown, of which:
3L2~i~;389
Figure 1 shows a perspective vie~ of one embodiment
of the automatic pipetting apparatus of the invention with
a sample plate secured thereto and its cover attached;
Figure 2 is a schematic illustration of the invention
sho~ing the functional relationship bet~een the sample
plate with its sample chambers, wash well, ~ast~ well and
a longitudinal application space with a microporous
support medium secured thereto beneath a pipette assembly
in which individual pipette barrels are moved as a unit up
and do~n and a plunger bar secured to plungers which are
moved up and down ~ith respect to the barrels;
Figure 2A schematically shows another embodiment of
the invention where the mounting plate includes two
stationary units and the pipette frame is longitudinally
movable with respect to the plate;
Figure 2B i8 a perspective view of another embodiment
of the sample plate further including a ro~ of dilution
wells;
Figure 3 i8 a side view partially cut a~ay and in
section ~ith the cover removed and taken from the view
along lines 3-3 of Figure l;
Figure 4 is a cross-section of the carriage and the
sample plate;
Figure 5 is a downward looking view along lines 5-5
of Figure 1 and shows the cross-section of the base at
level 5-5 and the track, carriage ~nd sample plates
beneath the pipetting assembly;
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Figure 6 is a cross-sectional vie~ taken along lines
6-6 of Figure 5 and sho~s the bas~, the carriage and
sample plate according to the invention and further sho~s
the translatio~ and guiding means by ~hich the track is
moved for~ard and back~ard ~ith respect to th~ pipetting
assembly;
Figure 7 is a for~ard looking vi~ taken along lines
7-7 of Figure 3 and sho~s partially broken a~ay, partially
cross-sectional parts of the pipette assembly slidably
mounted on posts secured to the base and including a
mounting plate ~hich moves up and do~n ~ith raspect to the
base and carrying a ro~ of barrels of individual pipettes
and an actuator plate movable vertically ~ith resp~ct to
the mounting plate for moving a plunger bar vertically for
moving individual plungers within the barrels of each of
the pipettes;
Figure 8 sho~s a top vie~ of the pipette assembly
showing in dashed lines the mounting platP of the pipette
assembly;
Figure 9 shows a cross-sectional vie~ taken along
lines 9-9 of Figure 7 and illustrates the relationship
bet~een the mounting plate, the tip bar and the plunger
bar and the means by ~hich the mounting plate is moved up
and do~n ~ith respect to the base;
Figure 10 is a similar vie~ to that of Figure 7 but
sho~s the mounting plate having been translated to a lo~er
position but ~ith the plunger bar remaining in an up~ard
1~13G3~9
-13-
position ~hereby the tips of the pipetters are in a
do~n~ard position, but the plungers are extended up~ardly
from each of the barrels of the pipettes
Figure 11 is a top cross-sectional vie~ taken along
lines 11-11 of Figure 10 and illustrates the motor and
rack and pinion system by ~hich the mounting plate is
moved up and do~n ~ith respect to the base;
Figure 12 is a cross-sectional vie~ taken along lines
12-12 of Figure 10 and illustrates the relationship of the
mounting bar after it has been moved do~n~ardly by the
rack and pinion system by operation of the motor turning
~ith re~pect to the frame mounted rack;as shown with Fig. 9;
Figure 13 is a vie~ similar to that of Figures 7 and
10 but illustrates the plunger actuator plate and plunger
bar moved do~nwardly with respect to the mounting plate
thereby forcing the plungers associated ~ith each of the
pipetters into their barrels and forcing any fluid
previously aspirated into the barrels out the tips of the
barrels;
Figure 14 shows a cross-sectional vie~ taken along
lines 14-14 of Figure 13 and illustrates the rack and
pinion system by ~hich the actuator plate and the plunger
bar secured thereto is moved up and do~n ~ith respect to
the mounting plate;
Figure lS is a cross-sectional vie~ taken along lines
15-15 of Figure 13 and illustrates the movement of the
actuator plate do~n~ardly ~ith respect to the mounting
389
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plate by operation o~ the rack ana pinion system
controlling relative movement bet~een the actuator plate
and the mounting plate; as shown with Fig. 9;
Figure 16 is a schematic illustration of the
microcomputer integrated circuit device receiving signals
from position detector circuits associated ~ith the
carriage, the mounting plate and actuator plate and
applying translation signals to motors for positioning the
carriage, the mounting plate and the plunger actuator
plate; and
Figure 17 is a functional flow chart illustrating the
stored program in the microcomputer device for
automatically washing, blotting, sampling and applying
samples to the support medium strip.
DESCRIPTION OF THE INVENTION
. _
Figure 1 illustrates in a perspective vie~ the
automatic pipetting apparatus 10 according to the
invention. The apparatus includes a base 12 on ~hich a
sample plate 16 is movably supported. The sample plate
includes a space 21 for securing a microporous support
medium 20 such as a cellulose acetate or agarose strip
used in the field of zone electrophoresis or other
separation techniques including the field of thin layer
chromatography. A cover 14 is provided behind ~hich a
pipette head 18 is sho~n.
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Figure 2 is a schematic illustration of th~ essential
mechanical elements of one embodiment of the invention
~ith the base and the cover removed. None of the mounting
apparatus is sho~n in Figure 2 so as to simplify the
explanation of the relationship of the sample plate 16 to
the pipette assembly 70. The translational means are
sho~n in a functional way rather than in actual mechanical
detail ~hich ~ill be sho~n in detail in the figures and
discussion below.
The sample plate 16 as sho~n includes a row of sample
chambers 24 as ~ell as a wash well 26, rinse or "waste~
well 28 and a longitudinal space 21 on which a microporous
support medium 20 is removably secured. The sample
chambers 24, the rinse ~ell 28 and the wash ~ell 26 are
provided on raised portions 22 of the sample plate. If
desired, a plastic cup may be provided in each of sample
chambers 24. A blotter space 32 between the rinse ~ell 28
and the sample chambers 24 is provided at substantially
the same vertical level 49 as the sample application space
21 on ~hlch the support medium 20 is secured.
As illustrated in Figure 2, the various regions of
the sample plate are longitudinally distinct, yet the
lateral spacings bet~een the sample chambers 24
corresponds to the application spots 19 on the support
medium 20 which is indicative of the fact that the barrels
92 of the pipette assembly 70 are arranged in a row
corresponding to the sample chambers 24. Liquid from
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~Z~3~3~
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those chambers is aspirated by the automatic pipetting
apparatus and is applied in a similar ro~ on the spots 19
of the microporous support medium 20.
It is advantageous to provide the sample chambers 24,
~aste ~ell 28 and wash ~ell 26 in raised portions 22 of
the sample plate 16 so that the mounting plate 80 of the
pipette assembly 70 need only go down to a common do~n~ard
position during all wash, ~aste, blot, ~ampling and
application operations. Ho~ever, it ~ould be obvious to
one of ordinary skill in the art that other arrangements
could be provided e~pecially ~here different levels of the
mounting plate could be prov~ded in the translation and
signalling apparatus for controlling the mounting plate
80. A detailed discussion of such translation and
signalling apparatus for controlling the mounting plate 80
i8 discussed belo~.
The schematic illustration of Figure 2 sho~s that the
sample plate 16 i~ tran~lated in for~ard and rearward
directions beneath the pipette assembly 70 by virtue of
the motor 40 turning a pinion 38 having its gears in
engagement with those of rack 36. As the shaft of the
motor 40 turns, the sample plate 16 carried by the rack 36
moves back and forth beneath the pipette assembly 70.
Turning no~ to the pipette assembly 70 sho~n in
Figure 2, a mounting plate 80 is translated up~ardly and
do~n~ardly by means of mounting plate motor 116 having its
pinion 120 engaging a rack 122. Thus, the entire mounting
'12~3~3F~9
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plate 80, and the microsyringe barrels 92 attached to the
barrel bar 88 which is secured to the mounting plate 80,
moves up and down in accordance with the turning of the
mounting plate motor 116. Similarly, the plungers 94
which are attached to plunger bar 90 and actuator plate 84
are moved up and do~n ~ith respect to mounting plate 80 by
operation of the turning of actuating plate motor 126 and
its pinion 130 engaging actuator rack 132. For purposes
of illustration, the actuator plate motor 126, its pinion
130 and the actuator plate rack 132 are sho~n on the
forward side of mounting plate 80, but the actual
apparatus illustrated in the subsequent figures is to the
rearward side of the mounting plate 80 through slots in
it.
Figure 2 therefore sho~s all of the essential
elements as far as the translation of the sample plate 16
backward and forward beneath the microsyringe barrels 92
and illustrates the upward and downward translation means
of the mounting plate 80 and the microsyringe barrels 92,
and the upward and downward motion of the plungers 94 and
the plunger bar 90 and actuator plate 84 with respect to
the mounting plate 80.
Figure 2A schematically illustrates an alternative
embodiment of the invention where the sample plate remains
immovable with respect to the base 12' ~ith the pipette
assembly 70' being mounted on rollers 300 for longitudinal
translation. Figure 2A illustrates that the sample plate
i~Z~3?39
-18-
may include two units, an application plate unit 16A and a
fluid plate unit 16B. The sample plate unit 16A is
adapted to removably secure a support medium 20', while
the fluid plate unit 1 6B includes a ro~ of sample chambers
24', a ~aste well 28', a ~ash well 26l and a blotting
space 32'. The operation of the alternat~ve embodiment is
similar to that of the embodiment of Figure 2 except ~hat
translation and signalling means are provided for
longitudinally translating pipette assembly 70' ~ith
respect to the sample plate(s) 16A, 16B. Details of such
translation and signalling means will be apparent to one
of ordinary skill in this art by virtue of the detailed
description of analagous translation and signalling means
described below.
Figure 2B illustrates an alternative sample plate 16'
~hich may include an additional ro~ of dilution wells 25
in addition to the row of sample chambers 24, waste ~ell
28, ~ash ~ell 26 and blotting space 32. Explanation of
the automatic diluting of sample fluid will be described
below with reference to Figure 2B.
Figure 3 is a side vie~ taken along lines 3-3 of
Figure 1 ~ith a portion of track 34 cut a~ay to show its
construction. The tracks 34 are supported by track
supports 48 which may also be seen in Figure 5. The
pipette assembly 70 is vertically supported from base
: mounting block 78 which is secured to the sides of the
. base 12 and is also further illustrated in Figure 5. The
lZt~ 9
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pipette assembly 70 includes a back plate 86 and a front
plate 73. One of the plurality of barrels 92 of the
pipette assembly is shown in an up~ard position.
A carriage 46 is slidably movably disposed on track
34 as more clearly seen in Figure 6. Racks 36 are secured
to carriage 46 and are movable with respect to the base 12
by means of the carriage motor 40 having its pinion 38 in
engage~ent with rack gear 36.
Notches are provided along the left edge of the
carriage 46. These notches cooperate ~ith a trip switch
to provide signals indicative of the longitudinal position
of the carriage. The wash notch 50, rinse notch 52, blot
notch 54, sample chamber notch 56, dilution chamber notch
57 (where the alternative sample plate 16' of Figure 2B is
used) and application notch 58 are illustrated in Figure
3.
Figure 4 illustrates a vertical cross-section through
the carriage 46 and the plate 16 and shows the actual ~ash
well 26, rinse well 28 and one of the sample cham~ers 24
on raised portion 22 of the plate 16. A blotter space 32
and a lateral application space 21 are illustrated on
plate 16. Blotter paper 30 is shown in blotter space 32
while a microporous support medium 20 such as cellulose
acetate or agarose is secured in lateral application space
21.
Figure 5 is a do~nward view taken along lines 5-5 of
Figure 1. Carriage 46 is shown supported by tracks 34 and
38~
~20~
movable in the rearward and for~ard directions by means of
motor 40, pinion 38 and rack 36 as also illustrated in
Figure 5. The sample plate 16 is disposed in a valley or
IJ-shaped cross-sectional structure 47 of carriage 46.
Position signalling notches on the sid~s of the
carriage 46 cooperate with trip s~itch 60 and trip switch
62 fixed to tracks 34. Spring loaded rollers 61 and 63,
respectively are forced against the longitudinal edges of
carriage 46 and into the notches as the carriage 46 moves
pa~t them. For example, the notches on the left hand side
46 include the wash notch 50, the ~aste notch 52, the hlot
notch 54, the sample chamber notch 56 and an application
notch 58. The notches correspond to the longitudinal
position of the wash ~ell 26, waste well 28, blotter paper
30, sample chambers 24 and application spots 19 when those
wells, chambers and blotting and application spots are
directly beneath the pipette barrels 92.
When the carriage 46 moves rearwardly ~here the wash
~ell 26 is directly beneath the barrels 92, the roller 61
moves into the ~ash notch 50 thereby tripping the trip
switch 60 for signalling the microprocessor associated
with electronic module 200 (Figures 3 and 16) that the
wash well is beneath the barrels 92. The trip switch 60
is likewise tripped when the roller 61 enters notches 52,
54, 56 and 58 to signal the position of the respective
other wells, chambers and spots beneath the barrels 92.
l Z~3B9
-21-
On the right hand side of the carriage 46 is notch 59
in ~hich the roller 63 is sho~n. A sample plate trip
s~itch 62 is thereby tripped to indicate that the carriage
is at its maximum forward position. Trip s~itch 65 is
mounted on the rear edge of carriage 46. Switch 65 closes
~hen the rear edge of the plate 16 is in position and
engages it thereby generating a signal that the plate 16
is properly in position on carriage 46. The stops 64
provide means for accurately longitudinally positioning
application spots 19 beneath barrels 92 ~hen carriage 46
is`in the maximum rear~ard position.
Also shown in Figure 5 is a start button 214 by which
the programmed microprocessor is signalled to start the
automatic sequence of events for the automatic pipetting
apparatus ~hich ~ill be explained in detail belo~. Lamp
212 provides a visual indication to the user of the
automatic pipetting apparatus 10 that the po~er is turned
on.
Figure 6 illustrates in a cross-sectional vie~ taken
along lines 6-6 of Figure 5 the means by ~hich the
carriage is translated ~ith respect to tracks 34. The
tracks 34 are supported upon base 12 by means of supports
48. The carriage 46 includes slots 149 in its sides on
~hich it slides on tracks 34.
Plate 16 as indicated above is disposed ~ithin a
notch or valley 47 of the carriage 46. The carriage
translation motor 40 is fixed to the base 12 and includes
~2 ~ 3
-22-
a shaft 42 ~hich is supported by means of shaft supports
44. Pinions 38 secured to shaft 42 have their gears in
engagement ~ith racks 36 ~hich are attached to the
carriage 46. As the motor 40 is turned in one direction
or the other under control of the microprocessor in the
electronic module 200 (Figure 16), the carriage 46 moves
in the for~ard or rear~ard directions.
Figure 7 illu~trates the pipette assembly 70 looking
rear~ardly along lines 7-7 of Figure 3. The mounting
blocks 76 are sho~n secured to the base mounting blocks 78
by means of scre~s 79. The mounting blocks 76 carry
vertical mounting posts 72 as illustrated in Figures 7, 10
and 13 and in the top view6 of Figures 8, 11 and 14. The
front plates 73 and back plate 86 are secured by means of
scre~s to mounting blocks 76.
A mounting plate 80 is vertically slidably supported
about the vert$cal mounting po~ts 72. Retainer bearings
74 provide sliding engagement bet~een the posts 72 and
vertical bearing blocks 75. The mounting plate 80 is
fastened to extension6 of bearing blocks 75 by means of
scre~s 81. By reference to Figures 7 and 13, it is seen
that mounting plate 80 may be moved from it~ up~srd
posit~on as sho~n in Figure 7 to its lo~er position as
sho~n in Figure 13 by its attachment to bearing blocks 75
and their sliding engagement on posts 72.
A barrel bar 88 i9 secured to mounting plate 80 by
means of ~cre~s 89. Mounted on barrel bar 88 are a
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plurality of pipette barrels 92 having their heads 193
secured ~ithin the barrel bar 88 in a manner to be
described below. As illustrated in the partial cut a~ay
of barrel bar 88, the barrel lock bar 91 secures the lower
portions of the barrels 92 to provide stability to the
barrels. Guide tips 97 include adjustable screws 95
extending below the bottom edge of the mounting plate 80
which cooperate ~ith the lower surfaces 49 of plate 16 to
accurately vertically position the lower tips 93 of
barrels 92 with respect to the support medium 20 and
blotting paper 30 disposed on lower surface 49 of plate
16. Such adjustment allows the droplets which form on the
ends of the tips 93, when plungers 94 are driven
do~nwardly within barrels 92, to "kiss" or be slightly
appl$ed either to the support medium or the blotting
paper. The droplets on the lower tips 93 of the barrels
92 are held because of their small size (as small as one
micro liter) and surface tension forces of the barrel
tips. When the tips are brought to a small distance
within the upper surface of the support medium 20 or
blotting paper 30, the droplets are relieved of the
surface tension holding them to their barrels and are
precisely applied to the blotting paper or to the support
medium.
Actuator guides 183 are secured to the mounting plate
80 and include grooves in which an actuator plate 84 is
inserted for sliding movement upwardly and downwardly with
lZ~63B9
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respect to the mounting plate 80. The actuator plate 84
has grooves in ~hich a plunger bar 90 is inserted. The
plungers 94 of the microsyringe barrels 92 are secured to
the plunger bar 90 and extend ~ithin the barrels 92. As
illustrated in Figure 7, the plungers 94 are at their
uppermost extent with respect to the barrels 92. The
actuator plate 84 is adapted to move do~n~ardly with
respect to the mounting plate 80, and through such action,
the plunger bar 90 moves do~nwardly ~ith respect to the
barrel bar 88 causing plungers 94 to move do~nwardly
within the barrels 92 thereby forcing any fluid ~ithin
such barrel~ out~ardly through the tips 93 of the barrels
and forming a droplet at the tips of the barrels.
Position B~ gnals are generated indicative of the
position of the mounting plate 80 with respect to the base
12 and the position of the actuator plate 84 and its
plungers 94 ~ith respect to the mounting plate 80. The
trip s~itch 106 mounted on the mounting plate 80
cooperates with the lower stop 115 mounted on the mounting
block 76 to provide a lo~er mounting plate position signal
when the mounting plate 80 reaches its lo~er extent. In a
similar way as shown on the right hand side of the
mounting plate 80, the upper trip switch 105 is mounted on
the mounting plate 80 and is shown tripped by contact ~ith
the upper stop 114. The upper trip s~itch 105 when
tripped provides a signal to the microprocessor of
lZ~63~9
-25-
elsctronics module 200 (Figure 3) indicative that the
mounting plate 80 is in its upper position.
The trip switch 108 mounted on left hand side
actuator 183 cooperates ~ith application position cam 100
and wash cam 102. The trip switch 108 is tripped by the
application cam 100 when the actuator plate 84 moves
upwardly with respect to the mounting plate 80 ana the
trip switch 108 is tripped by wash cam 102 as the actuator
plate 84 moves further upwardly. The do~n cam 104 trips
trip ~witch 110 mounted on the right hand side actuator
guide 183 when the actuator plate 84 reaches its maximum
downward travel where the plungers 94 are within the
barrels 92.
Figurè 9 is a cross-sectional view taken along lines
9-9 of Figure 7 and illustrates the mounting plate 80 and
the actuator plate 84 both in their upward positions.
Mounting plate motor 116 has driven the mounting plate 80
to the upward position by operation of pinion 120 on rack
122 which is secured to the back plate 86 and to the base
of the apparatus.
Turning no~ to Figure 10, the pipette assembly 70 is
shown with the unting plate 80 in the downward position
but the plunger bar 90 and the plungers 94 are in their
upward position ~ith respect to the barrel bar 88. The
barrels 92 are in a do~nward position in the wash well for
aspirating 5 ~1 fluid, for example, from the ~ash well 26.
.
:~2b~3~3~
-26-
Figure 11, a vie~ of the pipette assembly looking
down~ardly along lines 11~11 from Figure 10, illustrates
the drive mechanism by which the mounting plate 80 is
moved up and down ~ith respect to the base. A mounting
plate actuator motor 116 is fixed to the mounting plate 80
by means of a mounting screw 117. The output shaft of the
motor 116 has a gear 119 fixed to it. Gear 119 is engaged
with a left pinion gear 120L ~hich is mechanically coupled
to a right pinion gear 120R by meanæ of shaft 124. The
shaft is mounted to the mounting plate 80 by shaft
mountings 118. Racks 122, fixed to the back plate 86,
have their gears in engagement with pinion gears 120L and
120R. As the motor 116 turns in either direction under
microprocessor control, the mounting plate 80 is moved
upwardly or downwardly with respect to the base and the
back plate 86 by the rack and pinion mechanism. The front
view (Figure lO) of the pipette assembly 70 with the
mounting plate 80 in a downward position shows the racks
122 visible. The front view also illustrates, with the
mounting plate 80 in its maximum downward position, that
stop scre~s 95 of guide tips 97 are slightly above
surfaces 49 of the plate 16 indicative that tips 93 are
~lightly above the top edge 49 of the plate so that
droplets which form on the tips may l'kiss" the surface 49
and apply sample fluid to the support medium or blotting
paper.
:
12~3~9~
-27-
Figure 12 is a cross-sectional view of the pipette
apparatus taken along lines 12-12 of Figure 10 and sho~s
that the mounting plate actuator motor 116 has turned its
pinion gear 120 so that the mounting plate 80 has been
moved do~nwardly with respect to the base and the raised
portion 22 of the plate 16. Thus, the barrel 92 has been
lowered to be within a well of the raised portion 22 of
the sample plate. A wash well 26 is illustrated as an
example ~here the barrels 92 of the pipettes have been
lo~ered by the mounting plate 80 and where the fluid from
the wash ~ells have been aspirated into the barrels 92 by
virtue of the plungers 94 being pulled upwardly by means
of the actuator plate 84. It is apparent from Figures 11
and 12 that the mounting plate 80 is translated upwardly
and downwardly ~ith respect to the raised portion 22 by
means of the motor 116 turning and causing the pinion 120
to translate upwardly and down~ardly on fixed rack 122.
Turning no~ to Figure 13, the state of the pipette
apparatus 70 iB such that the actuator plate 84 has moved
do~n~ardly causing the plungers 94 to be inserted back
into the barrels 92 thereby positively displacing any
fluid ~h$ch has been aspirated ~ithin the barrels either
to an application space, a blotter, or to a ~aste ~ell.
It is seen that the trip switch 108 has been returned to a
condition such that any upward movement of the actuator
plate 84 will be tripped first by the application cam 100
and then th- wDsh cam 102 provlding D meanY for signa11ing
,'
-28-
the position of the actuator plate 84 ~ith respect to the
mounting plate 80.
Figure 14, is a downward looking vie~ along lines
14-14 of Figure 13 and illustrates the drive mechanism by
which the actuator plate 84 is translated up~ardly and
downwardly with respect to the mounting plate 80. A
plunger bar actuator plate motor 126 is fixed to the
mounting plate 80 by means of a mounting scre~ 127. The
motor 126 includes a gear 128 on its output shaft which is
in engagement with pinion gear 130R. Pinion gear 130R is
coupled to a pinion gear 130L by means of a shaft 134
which i~ supported by means of shaft mountings 118 which
also supports shaft 124 (see Figure 11). The actuator
plate 84 has actuator plate racks 132 fixed to the rear
side thereof which extend through slots 136 in the
mounting plate so as to engage the pini~ns 130L and 130R.
As the plunger bar actuator plate motor 126 i9 caused to
turn in either the clock~ise or the counterclockwise
direction, the actuator plate 84 is caused to move
upwardly or do~n~ardly with respect to the mounting plate
80. Figure 13 shows the slots 136 in the mounting plate
80 through which the actuator plate racks 132 extend.
Figure 14 also shows the means by which the barrel
heads 193 of the barrels 92 are removably fixed to the
barrel bar 88. The barrel bar 88 comprises a receiving
bar 138 having slots 140 provided along its front face.
The barrel heads 193 are inserted therein and secured by
~2~;3~39
--29--
means of a securing bar 139 which holds the barrels
vertically in place. The securing bar 139 is secured to
the receiving bar 138 by means of scre~s 141. The barrel
lock bar 91 is similarly constructed as the barrel bar 88.
The barrel bar 88 provida a removably securin~ means by
~hich the barrels 92 may be easily replaced due to ~ear or
breakage.
Figure 15 is a cross-sectional vie~ looking along
lines 15-15 of Figure 13 and illustrates the mounting
plate 80 in a do~nward pocition. $he actuator plate 84
has been tran~lated downwardly ~here the plunger bar 90 is
adjacent the barrel bar 88. Figure 15 illustrates the
actuator plate rackc 132 extending through slots of the
mounting plate 80 and their engagement with pinion gear
130 ~hich has been turned by means of the actuator plate
mntor 126. The barrel 92 is now in a rinse ~ell 28, for
example. The plunger 94 has been forced do~n by means of
the actuator plate 84 moving the plunger bar 90 to its
lo~ermost position. Of course, the ra$sed portion of the
plate 22 has moved longitudinally wlth respect to the
- pipette assembly bet~een the vie~s of Figures 12 and lS.
Figure 16 illustrates schematically the means by
which the carriage mounting plate and plunger actuator
bars are controlled to perform the automatic pipetting
operation. The dotted box 200 represents a microcomputer
integrated circuit device, preferably a microcircuit No.
, HD68POlV07 manufactured by the Hitachi Corporation. The
'~,' ~ ' ~ ' . - . .
. .
-
l~iG389
-30-
circuit includes a central processing unit 201A, a read
only memory 201B, a random access memory 201C, a timer
201D, an output interface circuit 201E and input interface
circuit 201F. The read only memory circuit 201B includes
stored soft~are by which the entire automatic operation is
controllea and will be discussed below.
Figure 16 illustrates the carriage motor 40, the
mounting plate motor 116 and the actuating plate motor 126
all under computer control via the motor driver circuits
202, 203, 204 such a6 circuits UDN-2952B manufactured by
the SPRAGUE Corporation. These motor driver circuits are
u6ed to control the speed of the motor and its direction
of rotation. Also provided in conjunction ~ith the motor
are electronic motor break circuits 205, 206, 207 which
are provided to quickly break the motor's rotation on
receipt of a translation signal by the computer 200. Such
electronic motor break circuits are preferably 2N6075
Triac circuits.
The po6ition detector circuits 208 represent the
circuitry with the trip switches 60 and 62 illustrated in
Figure 5 which indicate the position of the carriage 46
and the sample plate 16 ~ith respect to the pipette
assembly .
The position detector circuits 209 represent the
circuitry associated with the lower trip switch 106 and
the upper trip s~itch 105 ~hich signal the up~ard or
lZf~38~
-31-
do~n~ard limits of travel of the mounting plate 80 ~ith
respect to the base.
The position detector circuits 210 represent the
circuitry associated ~ith trip s~itches 108 and 110 ~hich
indicate the relative position of the actuator plate 84
~ith respect to the mounting plate 80. The signals
associated ~ith each of those position detector circuits
are represented as bei~g carried by a bundle of electrical
leads 215 to the input interface circuitry 201F of
10 electronic module 200.
The alarm circuit 211 i8 provided for the apparatus,
for example, such as a sounding device EAF14R06C
manufactured by Panasonic. Such circuit is act~vated and
a sound is generated to signal faults in the operation of
the apparatus or to signal the readiness of the machine.
Indicator circuit 212 represents an indicator lamp as
illustrated in Figure 5 to signal the user that the po~er
is on to the apparatus. The interlock circuit 213
represents the circuitry ~ith sample plate interlock trip
20 s~itch 65 ~hich indicates the presence or absence of the
sample plate on the carriage. Command circuit 214
represents a push button s~itch used to start or abort the
automatic pipetting application process.
In operation, the central processing unit 201A
receives the sequence of events instructions from the
s programs stored in the read only memory 20lB. The central
processing unit 201A then receives positional information
',
., .
,,
,~ ,
, .
3~9
-32-
concerning the moving mechanisms of the apparatus by means
of reading and decoding the binary coded data present at
the input interface 201F which receives information via
leads 215 from the position detector circuits 208, 209,
210.
The microproces~or CPU 201A then receives an input
command to start or abort the process by means of reading
and decoding the binary coded data present at the input
interface 201F which is connected to the command circuit
214 whlch may be the push button 214 illustrated in Figure
5.
Figure 17 illustrates in flo~ chart form the
operations of the CPU 201A under program control. The CPU
201A determines the validity of a command to start the
processing by means of reading and decoding the binary
coded data pre~ent at the input interface 201F which
receives a signal from the interlock circuit 213. This
operation insures that the plate 16 i8 fully inserted into
; the carriage.
The CPU 201A causes the motors 40, 116 or 126 to turn
in the required direction be means of ~riting the
appropriate binary coded data to the output interface
circuit 201E which is connected to the motor drive
circuit~ 202, 203 and 204. The microprocessor CPU 201A
then causes the mechanism movement to stop precisely when
the re~uired location is reached by writing the
appropriate binary coded data to the output interface
, . . ..... ..
lZ~i3~
-33-
circuit 201E ~hich is connected to the motor drive
circuits 202, 203, 204 to aisable the drive and then
~riting the appropriate binary coded data to the output
interface 201E ~hich is connected to the motor break
circuits 205, 206, 207 to apply electronic breaking.
The microprocessor circuit 201A then signals that the
pipetting apparatus is ready or that a plate has been
completed or that a failure has occurred by means of
writing the appropriate binary coded data to the output
interface circuit 201E connected to the alarm circuit 211
to sound an alarm.
The timer 201D of Figure 16 is used by the
m$croproce~sor CPU 201A to determine eleatrical or
mechanical failures of the positioning mechanism. This is
accomplished by means of measuring the elapsed time during
a command to drive any motor. If the event is not
completed ~ithin the prescribed length of time, the drive
command i6 aborted and the alarm is activated by means of
the microproce~or CPU 201A writing the appropriate binary
coded data to the output interface connected to the alarm
circuit 211. The timer 201D i8 also used to determine the
repetition rate of the alarm thereby allowing the
microprocessor circuit CPU 201A to encript and communicate
to the operator the nature of the failure.
As sho~n in Figure 17, once the automatic pipetting
apparatus of the invention is running, a ~ide variety of
... .
dlffer-nt applic-tionc may be achieved. The ~equence of
'
~.
.',
lZt~638~
-34-
operations sho~n in Figure 17 is preerred in that first,
five microliters of cleansing agent such as distilled
water is aspirated into the barrels of the pipettes from
the wash well. Next, the water in the barrels is
dispensed into the waste well. Then the applicator tips
are dried by lowering them to a blotter pad as illustrated
in Figure 5.
Next, the barrels are moved to their upward position
with the plungers in their do~nward position, the carriage
0 i5 moved rearward and the barrels are lowered into the
sample chambers 24. The plungers are raissd thereby
dra~ing a small amount of each sample of liquid, for
example, patient blood to be tested. Where no dilution of
the blood samples is desired, the samples are applied
precisely to the cellulose acetate or agarose strip. The
barrels are raised again. Again, the carriage is moved
forward until the wash well is beneath the barrels and the
mounting plate is lo~ered such that distilled ~ater is
again dra~n into the barrels and then dispensed into the
~0 waste well.
Where dilution of the sample liquids is desired, the
sample plate of Figure 2B may be substituted for that of
Figure 2 and the computer program illustrated by the flo~
chart of Figure 17 branches to the dilute routine. A
preferred routine for diluting the samples is to dra~ an
additional four micro liters of fluid from the wash well.
This action results in each of the barrels being filled
:
: , .
~2~63f~9
-35-
~ith four micro liters of diluting fluid (e.g, ~ater) and
one micro liter of blood (or other liquid) sample. Next
the entire five micro liters of fluid of each barrel is
applied to the respective dilution chambers of the
dilution ro~ of the plate of Figure 2B. This process may
be repeated a desired number of times to effect mixing of
the sample ~ith the dilution fluid (~ater). Finally a one
micro liter sample of the diluted blood sample is drawn
into each barrel according to the description presented
previously. The routine then proceeds as described above
~here the one micro liter of diluted liquid sample is
applied to the support medium.
The dilution routine described above is preferred,
but other routines may be used to effect good mixing of
the blood sample with diluting liquid. For example, a
first predetermined amount of liquid sample in each barrel
may be applied to the dilution ~ells. A predetermined
amount of ~ash liquid may then be applied to the dilution
~ells. After mixing the combination of the ~ash liquid
and liquid sample of the dilution ~ells, (for example by
the mixing technique described above), a small amount of
liquid samples is aspirated from the dilution ~ells and
applied to the support medium.
An appendix to this specification is filed herewith
and the appendix is to be retained on the Patent file and
includes a source listing of the computer program written
in HD68POlV07 Hitachi Assembly language which is stored
in the read only memory 201B so as to automatically
control the pipetting,
2 ~ 3
-36-
cleansing, blotting, diluting ~at the operator's option)
and other functions described above.
Various modifications and alterations in the
described structures ~ill be apparent to those skilled in
the art of the foregoing description ~hich does not depart
from the spirit of the invention. For this reason, these
changes are desired to be included in the appended claims.
The appended claims recite the only limitation to the
present invention and the descriptive manner ~hich is
employed for setting forth the embodiments and is to be
interpreted as illustrative and not limitative.
12~36389
_ APPENDIX TO SPECIF1 ~TION
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