U.S. patent application number 10/056511 was filed with the patent office on 2002-12-05 for chemical dispensing system and method.
Invention is credited to Aghassi, Nora B., Ardi, Paul J., Franceschini, Kim G..
Application Number | 20020182115 10/056511 |
Document ID | / |
Family ID | 25293042 |
Filed Date | 2002-12-05 |
United States Patent
Application |
20020182115 |
Kind Code |
A1 |
Aghassi, Nora B. ; et
al. |
December 5, 2002 |
Chemical dispensing system and method
Abstract
A system and method for applying one or more chemicals to a
tissue sample is provided. The system preferably includes a
cassette for housing a slide device, a film, and an injection
system. The slide device preferably includes a specimen slide for
containing the tissue sample and a cover plate connected to the
specimen slide. The film is preferably moveable through the
cassette along guide rollers and preferably contains a plurality of
containers containing one or more chemicals. The injection system
may include a piston for displacing the chemicals from the
containers through an injection port to the tissue sample. The
cassette may be placed on a cassette driver that contains a
motor-driven shaft for driving the piston and moving the film.
Inventors: |
Aghassi, Nora B.; (Hot
Springs, AZ) ; Franceschini, Kim G.; (Austin, TX)
; Ardi, Paul J.; (Hot Springs, AZ) |
Correspondence
Address: |
JACKSON WALKER LLP
Suite 2100
112 E. Pecan
San Antonio
TX
78205-3731
US
|
Family ID: |
25293042 |
Appl. No.: |
10/056511 |
Filed: |
January 31, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
10056511 |
Jan 31, 2002 |
|
|
|
08844553 |
Apr 18, 1997 |
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Current U.S.
Class: |
422/400 |
Current CPC
Class: |
G01N 35/00009 20130101;
Y10T 436/112499 20150115; Y10T 436/2575 20150115; G01N 1/312
20130101 |
Class at
Publication: |
422/100 |
International
Class: |
B01L 003/02 |
Claims
I claim:
1. A system for applying a chemical to a tissue sample, comprising:
a cassette; a slide device for holding the tissue sample, the slide
device being adapted to be housed within the cassette during use
and comprising a specimen slide and a cover plate, the cover plate
being attachable to the specimen slide such that a head space
exists therebetween, the slide device farther comprising an
injection port communicating with the head space; and a film
adapted to be located within the cassette for delivering a chemical
to the injection port, the film comprising a container for holding
the chemical.
2. The system as recited in claim 1, wherein the cassette further
comprises an open end for receiving the slide device.
3. The system as recited in claim 1, wherein the slide device
comprises a spacer for defining the head space, the spacer being
disposed between the specimen slide and the cover plate, the spacer
being located around outer edges of the specimen slide and the
cover plate.
4. The system as recited in claim 1, wherein the cassette comprises
(1) a cavity being adapted to house the cover plate and (2) a
raised protrusion on a side of the cavity, and further comprising a
spacer being disposed between the raised protrusion and the
specimen slide such that the specimen slide is a pre-determined
distance from the cover plate, the cover plate being within the
cavity and being unconnected to the specimen slide.
5. The system as recited in claim 1, wherein the cassette further
comprises a cavity, the cavity being adapted to house the slide
device.
6. The system as recited in claim 1, wherein the cassette further
comprises a cavity and a removable cap adapted to cover at least a
portion of the cavity.
7. The system as recited in claim 1, wherein the specimen slide and
the cover plate are adapted to be snapped together.
8. The system as recited in claim 1, further comprising a slide
holder for attaching the specimen slide to the cover plate.
9. The system as recited in claim 1, wherein the head space is
between about 0.0005 inch and about 0.004 inch.
10. The system as recited in claim 1, wherein the head space is
between about 0.0015 inch and about 0.0025 inch.
11. The system as recited in claim 1, wherein the slide device
further comprises a first end, a second end, and a relief port, the
first end being opposite to the second end, and wherein the
injection port communicates with the head space at the first end,
and wherein the relief port communicates with the head space at the
second end.
12. The system as recited in claim 1, wherein the slide device
further comprises a first end, a second end, and a relief port, and
wherein the injection port communicates with the head space at the
first end, and wherein the injection port comprises a conduit, the
conduit communicating with the head space to allow the chemical to
pass through the conduit into the head space.
13. The system as recited in claim 1, wherein the film comprises a
plurality of holders for holding chemically filled containers.
14. The system as recited in claim 1, wherein the container
comprises a cavity for holding the chemical, a plurality of
protrusions extending from an inner surface of the cavity, and a
cap for sealing the cavity, the cap comprising a plurality of
complementary protrusions adapted to mate with the protrusions of
the cavity.
15. The system as recited in claim 1, wherein the container
comprises a cavity for holding the chemical and a removable cap for
sealing the cavity, the cap comprising: (a) a slit terminating in a
wall, the slit being adapted to deliver the chemical from the
cavity to the injection port; and (b) a plurality of protrusions
extending from an outer surface of the cap.
16. The system as recited in claim 1, wherein the container
comprises (a) a cavity for holding the chemical; (b) a plurality of
protrusions extending from an inner surface of the cavity; (c) and
a cap shaped to mate with a portion of the cavity for sealing the
cavity, the cap comprising a plurality of complementary protrusions
adapted to be inserted between the protrusions of the cavity for
locking the cap within the cavity.
17. The system as recited in claim 1, wherein the film extends
along an edge of the cassette and forms a substantially
horseshoe-shaped loop at an interior portion of the cassette.
18. The system as recited in claim 1, wherein the container
comprises a wall to separate two or more chemicals within the
container.
19. The system as recited in claim 1, further comprising a guide
roller located within the cassette, the guide roller engaging the
film and being adapted to rotate to move the film during use.
20. The system as recited in claim 1, further comprising a waste
tank containing absorbent material for collecting chemical that
exists the slide device, the waste tank being connected to an end
of the slide device.
Description
BACKGROUND OF THE INVENTION
[0001] 1. Field of the Invention
[0002] This present invention relates to histological and molecular
pathology and more specifically to an immunohistochemistry staining
system for staining tissue samples with a variety of chemicals to
facilitate examination of the samples.
[0003] 2. Description of the Related Art
[0004] Staining tissue with chemicals is generally well known in
the field of molecular pathology. Tissue samples may be subjected
to histological and molecular pathological tests including in-situ
hybridization, polymerase, and to chemical stains which are
commonly referred to as "special stains". A variety of chemicals
may be used in the staining process, including biological reagents,
antibodies, buffers, and deionized water. After a tissue is stained
and rinsed with certain chemicals, one skilled in the art can
identify the tissue type and any abnormalities in the tissue. Such
methods may be used to identify various diseases in the tissue.
Several methods of staining tissue are currently in use.
[0005] One conventional method utilizes capillary action in which a
chemical in liquid state is drawn up a narrow space between two
slides due to the attraction between the molecules of the chemical
and the slides. One of the slides contains a tissue sample to be
contacted with the chemical. An operator is generally required to
pipette the appropriate staining chemicals into containers into
which the slides are placed. Filling the containers tends to
require a substantial amount of time. The slide pair is then
manually or mechanically inserted vertically into each container in
an order dependent on the type of chemical in each container.
Manually placing the slide pair in the correct container may be
difficult if the containers are not labeled for the appropriate
chemical. The method may be also be difficult when performed
mechanically if a mechanical arm is not properly programmed to move
the slide pair sequentially to the proper containers. A computer
may be used to program the movements of a mechanical arm. The
length of time that the slide pair is in each container is
determined by using a watch or a computer.
[0006] Another method for staining tissue uses an automated
dispensing head or nozzle to drop or spray chemicals onto a slide's
upper surface which contains the tissue sample. This may also be
performed manually. An evaporation inhibitor is placed over the
slide after a chemical agent is placed onto the tissue sample. The
movable dispensing head draws the desired chemical from a container
and dispenses the chemical on top of the appropriate slide. One
limitation of this method is that a drop of liquid may form at the
tip of the head after each dispensing operation and may drop onto
another slide as the head moves. Also, the containers need to be
filled with chemicals before they are dispensed, which requires
labor. The length of time that elapses before each chemical is
washed from a slide may be controlled by a computer.
[0007] A similar method involves dispensing a chemical from a
dispenser pre-packaged with the chemical so that a person does not
have to fill the dispenser. After each dispensing operation, the
dispenser withdraws the drop remaining on the tip of its nozzle to
prevent the drop from falling on a slide. The dispenser is placed
in a rotating tray located above a slide tray. The tray moves the
dispenser above the slide on which a chemical is dispensed. A
computer is used to control the movement of the tray and to
determine the length of time before a chemical is washed from a
slide.
[0008] U.S. Pat. No. 5,232,664 relates to a dispenser for
delivering small amounts of liquid to a sample. The dispenser
includes a reservoir chamber linked to a dispense chamber. The
dispense chamber communicates with a nozzle through an outlet line.
A number of such dispensers are positionable within a reagent tray
that is rotated by a drive carousel to perform an immunoassay. This
patent is incorporated by reference as if fully set forth
herein.
[0009] Turning to another method, chemicals may also be dispensed
manually or from a mechanical transfer head into a gap between a
cover plate and a slide containing a tissue sample. The slide and
cover plate are in vertical positions. Gravity forces the chemicals
through the gap. The surface tension between the slides prevents
chemicals from flowing immediately out of the bottom of the gap.
The transfer head may be filled with the appropriate chemical after
each dispensing. Such a method requires significant time and labor.
A computer may be used to control the movements of the transfer
head and to time how long a slide is exposed to each chemical.
[0010] The methods presented above tend to be problematic. Some of
the methods involve steps that are manually performed and require a
significant amount of time. For example, an operator may be
required to pipette a chemical onto a slide and/or fill chemical
containers used for staining tissue. The operator may also have to
carefully label these containers and time how long each chemical
contacts the tissue. Such processes are labor-intensive and may be
subject to significant variation due to human error. Further, some
of the methods require the use of expensive automated equipment and
a computer to control the equipment.
[0011] It is therefore desirable that an improved tissue staining
system be derived which is less labor intensive. Further, it is
desirable that the system requires less expensive equipment to
stain the tissue. A system with these features would tend to
increase the quality of the resulting stained tissue while reducing
operating costs.
SUMMARY OF THE INVENTION
[0012] The problems outlined above are in large part solved by an
improved tissue staining system and method of the present
invention. That is, the system hereof does not require expensive
equipment such as a computer in order to function. Further, the
system may be automated such that less labor is needed to operate
the system.
[0013] An embodiment of the system includes a cassette for housing
the staining system. The cassette preferably has an open end into
which a removable slide device may be placed. The slide device
preferably includes a specimen slide for holding a tissue sample
and a cover plate located above the specimen slide. A film is
preferably located within the cassette for delivering one or more
chemicals to the slide device. As described herein, a "chemical" is
taken to mean any substance added to the tissue sample to
facilitate testing or examination of the tissue sample, is
including but not limited to a biological reagent, an antibody, a
buffer, a label, a chromogen, a solvent, a resin and/or deionized
water. Each chemical may be reactive or unreactive with the tissue
sample.
[0014] The specimen slide and cover plate are preferably attached
together in spaced relation to form a head space therebetween. A
plastic (e.g., mylar) spacer may be placed between the cover plate
and specimen slide. This spacer preferably creates the head space
and preferably seals the outer edges of the slide device. The slide
device may be held together with a slide holder. Alternately, the
specimen slide and the cover plate may be constructed such that
they can be snapped together to form a fixable engagement On
opposite ends of the slide device, an injection port and a relief
port preferably communicate with the head space formed within the
slide device. The injection port preferably includes a conduit, and
that conduit preferably has a pointed end. A chemical may be passed
through the injection port into the head space and then out of the
head space through the relief port.
[0015] The slide device may be placed into the open end of the
cassette. An injection piston preferably located within the
cassette adjacent to the injection port may reciprocate in a
direction toward the injection port. This reciprocating motion
preferably enables the injection piston to contact containers
disposed on the film. The containers contain one or more chemicals
to be applied to the tissue sample, and holders may be used to
attach the containers to the film. The film preferably extends
along the edge of the cassette and forms a loop (e.g., in a
horseshoe shape) in the interior of the cassette. Guide rollers
within the cassette preferably move the film through the cassette
such that the containers disposed within the film are passed to a
location between the injection piston and the injection port.
[0016] The reciprocating motion of the injection piston is
preferably synchronized with the movement of the film such that the
piston contacts each holder of the film. In the case that a
container is disposed within the holder contacted by the piston,
the piston preferably forces the container against the pointed
conduit end within the injection port. The pointed conduit end
preferably punctures the container. The container preferably
ruptures, causing the chemical(s) within the container to be
released into the injection port. The piston preferably creates
pressure within the injection port to cause the released
chemical(s) to be positively displaced through the injection port
and into the head space to stain the tissue sample. In the case
that the head space contains a chemical that has been previously
injected, such previously injected chemical is preferably displaced
out of the head space by the pressure derived from the piston. The
displaced waste chemical is preferably passed through the relief
port and into a waste tank. The waste tank preferably contains
absorbent material to facilitate collection of such waste
chemicals.
[0017] The holders of the containers are preferably spaced apart by
equal distances along the length of the film. The contact time
between a chemical and a tissue sample is preferably determined by
the distance between containers on the film. A specified number of
holders may be left empty to create a predetermined amount of time
between (a) the injection of a first chemical into the head space
and (b) the injection of a second chemical into the head space and
the simultaneous displacement of the first chemical from the head
space. The motion of the film and the displacement of the injection
piston are preferably synchronized such that each holder in the
film is contacted by the injection piston.
[0018] The containers may hold more than one chemical. In an
embodiment, two or more chemicals within the containers are
separated by an interior wall within the container to prevent the
chemicals from mixing before injection. When a container holding
more than one chemical is forced against the pointed conduit end,
each of the wall segments within the container is preferably
punctured to allow the chemicals to mix before being injected into
the slide device.
[0019] In an embodiment, the cassette may be placed onto a cassette
driver for driving the guide rollers and the injection piston. The
cassette driver preferably includes a shaft which may extend from
the top of the cassette driver into the bottom of the cassette. The
shaft is preferably a spline joint shaft which may be connected to
the guide rollers and the piston. A gear drive motor preferably
rotates the shaft at a constant speed, causing the guide rollers
and the piston to move at a constant speed.
[0020] The cassette driver also preferably includes a heat pad and
a thermostat for controlling the temperature of the chemicals in
the head space of the slide device. The heat pad may be used to
heat or cool the tissue sample and chemicals in the head space. The
thermostat may maintain the heat pad within a predetermined
temperature range or it may cause pulse changes in the temperature
of the heat pad. In this way, the tissue and chemicals may be
subjected to any predefined temperature profile. Maintaining the
temperature of the tissue sample and chemicals within a selected
range may facilitate or control reaction of the injected chemicals
with the tissue sample. The cassette may be engaged with the
cassette driver such that the heat pad is positioned under the
slide device. The cassette driver preferably contains a pressure
sensitive switch enabling automatic activation of the cassette
driver motor when the cassette is detected on top of the cassette
driver. A pre-set timing device may be used to stop the motor after
a predetermined amount of chemicals have contacted the tissue
sample.
[0021] In an embodiment, the tissue staining system includes a
plurality of cassette drivers and a control panel for individually
operating each of the cassette drivers. The control panel is
preferably coupled to the motor and heat pad of each of the
cassette drivers. A plurality of cassettes may be placed on the
cassette drivers to allow a plurality of tissue samples to be
stained simultaneously.
[0022] In an alternate embodiment, a portable cassette includes a
battery operated solenoid piston for injecting chemicals into a
slide device. The slide device is preferably located proximate the
center of the cassette. The portable cassette preferably includes a
twin drive motor for rotating guide rollers to move a film. The
film preferably delivers containers to the solenoid piston. The
portable cassette preferably includes a heat pad located under the
slide device and a waste tank for collecting chemicals which are
displaced from the slide device. The portable cassette may include
printed contacts for activating the heat pad and/or the solenoid
piston.
[0023] Each of the embodiments discussed above may be combined or
used individually.
[0024] Further advantages of the present invention will become
apparent to those skilled in the art with the benefit of the
following detailed description of the preferred embodiments and
upon reference to the accompanying drawings.
BRIEF DESCRIPTION OF THE DRAWINGS
[0025] FIG. 1a is a perspective view of a cassette, a slide device,
and a slide holder which are part of a tissue staining system.
[0026] FIG. 1b depicts a cross-section of the slide device.
[0027] FIG. 1c depicts a side view of an alternate embodiment of
the slide device.
[0028] FIG. 2 is a perspective view of the cassette depicting the
slide device engaged within the cassette.
[0029] FIG. 3a is a top plan view of an interior portion of the
cassette.
[0030] FIG. 3b is a side view of the interior portion of the
cassette.
[0031] FIG. 3c is a cross-sectional view along plane II of FIG.
3a
[0032] FIG. 3d is a cross-sectional view along plane 111 of FIG.
3a
[0033] FIG. 4a is a top plan view of an injection end of the slide
device and a portion of a film.
[0034] FIG. 4b is a front view of a portion of the film.
[0035] FIG. 5 is a perspective view of a cassette driver.
[0036] FIG. 6 is a perspective view of a multiple cassette driver
unit.
[0037] FIG. 7 is a perspective view of a multiple cassette driver
system.
[0038] FIG. 8 is a perspective view of a portable cassette.
[0039] FIG. 9 is a cross-sectional view of a portion of a tissue
staining system, including a cassette and a cassette driver.
[0040] FIG. 10 is a cross-sectional view of a tissue staining
system, including a cassette and a cassette driver.
[0041] FIG. 11 is a cross-sectional view of the locking mechanism
of a chemical container which belongs to a tissue staining
system.
[0042] FIG. 12a is a perspective view of a portion of a cassette
and a portion of a film belonging to a tissue staining system.
[0043] FIG. 12b is a cross-sectional view of a portion of a film
belonging to a tissue staining system.
[0044] While the invention is susceptible to various modifications
and alternative forms, specific embodiments thereof are shown by
way of example in the drawings and will herein be described in
detail. It should be understood, however, that the drawings and
detailed description thereto are not intended to limit the
invention to the particular form disclosed, but on the contrary,
the intention is to cover all modifications, equivalents and
alternatives falling within the spirit and scope of the present
invention as defined by the appended claims.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0045] Turning now to one embodiment of the present invention, FIG.
1 illustrates an outside perspective view of cassette 1 which is
part of an improved tissue staining system. As described herein,
"cassette" is taken to include any housing for containing a tissue
sample to facilitate the application of chemicals onto the tissue
sample. The cassette in FIG. 1 is substantially rectangular and
contains an optional protective covering on its top surface that
covers the interior portion of the cassette. It is to be
appreciated that the cassette could have any of a variety of shapes
(e.g., circular, triangular, etc.). Cassette 1 preferably contains
a film access door 2 that may be located on the top, bottom, or
side of the cassette. Cassette 1 preferably further includes a
container access door 3 to allow access to an interior portion of
the cassette.
[0046] FIG. 1a illustrates slide device 5 into which a tissue
sample 6 may be placed. Slide device 5 preferably includes a
specimen slide 9 and a cover plate 8. A spacer 10a is preferably
placed between specimen slide 9 and cover plate 8. The spacer may
be made of a plastic such as mylar. Spacer 10a preferably extends
along the length of slide device 5 and seals the outer edges of the
slide device to maintain chemical between the specimen slide and
the cover plate. A slide holder 7 preferably holds slide device 5
together by clamping specimen slide 9 to cover plate 8 at one end
of slide device 5. An open end 4 of cassette 1 is preferably shaped
to receive slide device 5.
[0047] FIG. 1b depicts a cross-section of slide device 5 taken
along plane I. Cover plate 8 is preferably located above specimen
slide 9. Spacer 10a is preferably located between cover plate 8 and
specimen slide 9 at their outer edges. The thickness of spacer 10a
is preferably in the range between about 0.0005 inch and about
0.004 inch, more preferably between about 0.001 inch and about
0.002 inch, and even more preferably about 0.0015 inch. The
thickness of spacer 10a preferably creates a head space 10b between
cover plate 8 and specimen slide 9. The head space between cover
plate 8 and specimen slide 9 preferably has a width that is the
same as the thickness of spacer 10a. When chemicals are injected
into slide device 5, they preferably fill head space 10b and
interact with tissue sample 6. Spacer 10a may be disposed about the
entire perimeter of the slide device to surround the head
space.
[0048] In an alternate embodiment, specimen slide 9 and cover plate
8 are capable of being attached in fixable engagement without slide
holder 7. The specimen slide and cover plate may be snapped
together. Alternately, the cover plate may contain substantially
L-shaped members 50 as depicted in FIG. 1c, which depicts a cross
sectional view of slide device 5. The L-shaped members serve as a
base for supporting specimen slide 9, which may be slid into
engagement with cover plate 8. The tolerance between the specimen
slide and the cover plate is preferably sufficiently low to prevent
liquid chemical from exiting head space 10 through the outer edges
of the slide device.
[0049] FIG. 2 illustrates slide device 5 positioned within open end
4. A waste tank 11a is preferably placed in open end 4 near the end
of slide device 5. The waste tank may be used to collect waste
chemical that exits head space 10b. The waste tank is preferably
detachable from the end of slide device 5 to allow it to be removed
and replaced. In an embodiment, the waste tank contains absorbent
material to facilitate removal of the waste chemical. Preferably,
this absorbent material is an absorbent non-woven structure.
[0050] FIG. 3a depicts a top plan view of an interior portion of
cassette 1. Slide device 5 and waste tank 11 are preferably located
in open end 4 of cassette 1 during the tissue staining process. A
film 12 is preferably used to deliver chemicals to slide device 5
for application onto the tissue sample. The film is preferably
substantially flexible and may be a polymeric film, a ribbon film,
or any belt-like member that is sufficiently flexible to be drawn
around guide rollers 14 and 15 and moved through cassette 1. Film
12 may be loaded into and removed from the cassette through film
access door 2. Film 12 preferably extends about the perimeter of
cassette 1 and may form a horseshoe-shaped loop within the interior
portion of the cassette.
[0051] Guide rollers 14 and 15 may contain sprockets and preferably
engage film 12. The guide rollers are preferably connected to shaft
18 via secondary shafts 58. Shaft 18 is preferably a splined shaft
that is rotatable to cause guide rollers 14 to rotate. Rotation of
the guide rollers preferably causes movement of the film throughout
the cassette. A plurality of containers 13a are preferably disposed
on the film. Each container 13a preferably contains between about
50 microliters and about 200 microliters of chemical, and more
preferably between about 100 microliters and about 150 microliters
of chemical.
[0052] Guide rollers 15 preferably engage film 12 and rotate to
move the film about the cassette and through the horseshoe-shaped
loop depicted in FIG. 3a. The cassette preferably includes an
injection system for injecting chemicals into injection port 17.
The injection system preferably includes an injection piston 16
located near an end of slide device 5. Piston 16 is preferably
attached to rotating shaft 18 via cam 59 which displaces the piston
in a direction toward the film 12 at a location proximate the apex
of the horseshoe-shaped loop. Piston 16 preferably reciprocates
along an imaginary axis that extends longitudinally through
injection port 17. Piston 16 preferably contacts container 19 and
ruptures the container to release chemical into injection port 17.
Injection port 17 and relief port 21 preferably communicate with
the head space and may be disposed on opposite ends of the slide
device. The piston preferably creates pressure within injection
port 17 to force the chemical through the injection port and into
the head space 10b where it contacts the tissue sample.
[0053] FIG. 4a depicts a top plan view of conduit 22 which
preferably forms a portion of injection port 17 of slide device 5.
Conduit 22 preferably includes a sharp or pointed end for
puncturing containers as depicted in FIG. 4a The piston preferably
contacts the containers and forces them against hollow conduit 22.
The containers preferably rupture as a result of the contact with
conduit 22 and piston 16 and the chemical within the ruptured
container is preferably directed through conduit 22 into the head
space under pressure derived from the reciprocating motion of the
piston. Film 12 preferably carries container 13a to a location
adjacent to the sharp end of conduit 22. The film is preferably
sufficiently flexible to bend to allow container 13a to be pressed
against the end of conduit 22.
[0054] In an embodiment, container 13a contains two or more
chemicals to be injected into slide device 5. In some cases it may
be necessary to simultaneously apply two or more chemicals to the
tissue sample without mixing the chemicals beforehand. Container
13a may contain an interior wall having one or more wall segments
13c for compartmentalizing the container to prevent the mixing of
two or more chemicals within the container prior to injection. Wall
13c is preferably punctured by conduit 22 during injection to allow
the chemicals to mix as they are injected into slide device 5. The
chemicals are preferably positively displaced from container 13a
into head space 10b where they contact the tissue sample. In the
case that a first chemical resides in head space 10b at the time a
second chemical is injected into container 13a, the first chemical
is preferably positively displaced through relief port 21 out of
head space 10b by the second chemical and/or the pressure within
the head space derived from piston 16.
[0055] FIG. 3b depicts a side view of cassette 1. Film 12
preferably includes a plurality of holders 13b for attaching
containers 13a to the film. Holders 13b of film 12 are preferably
spaced apart by equal distances. The containers 13a are preferably
removable and replaceable on the film to allow the film to be
reused after the tissue staining process has been completed.
Container access door 3 preferably enables access to an interior
portion of the cassette to allow containers to be placed into empty
holders of the film after the film has been loaded into the
cassette. In this manner, user-specified chemicals may be added to
the film to be injected into slide device 5 to allow for customized
staining protocols with respect to selection of the applied
chemical.
[0056] The time of contact between a chemical and a tissue sample
may be predetermined by the spacing between containers on the film.
Selected holders may be left empty to increase the spacing between
adjacent containers, thereby lengthening the contact time between a
given chemical and the tissue sample. In operation, the
reciprocation of the piston and the movement of the film preferably
occur at constant speeds and are synchronized such that the piston
contacts each holder on the film.
[0057] FIG. 3c further illustrates a cross-section of cassette 1
along plane II of FIG. 3a, depicting guide rollers 15 and an end of
slide device 5. FIG. 3d is a cross-sectional view along plane III
of FIG. 3a. A heat pad 20 is preferably located under slide device
5 to control the temperature of chemicals and tissue sample within
head space 10b. Although heat pad 20 is shown in FIG. 3a to extend
through an opening in base 52, it also may reside entirely within
cassette 1 between base 52 and slide device 5. Heat pad 20 may be
used to heat or cool the chemicals and tissue sample to control any
reaction that may occur between the chemicals and the tissue
sample. The temperature of the heat pad 20 is preferably controlled
to maintain the temperature of the tissue sample and chemical
within a predetermined range. The temperature of the heat pad may
be continuously adjusted as a function of the type and/or amount of
chemical(s) injected into the head space. Heat pad 20 may be any
conventional heat exchange of thermoelectric device. Heating coils
or conduits adapted to contain a heat transfer medium may be
contained within heat pad 20 to control the temperature at its
surface.
[0058] Waste tank 11a is preferably placed at the end of slide
device 5 during the tissue staining process. An absorbent material
11b is preferably located in the bottom of waste tank 11a The
absorbent material 11b preferably absorbs waste chemical passing
from relief port 21 and facilitates the disposal of the waste
chemical.
[0059] FIG. 4b shows a side view of a portion of film 12 which
preferably contains openings 23. Each of the guide rollers may
contain teeth 54 (shown in FIG. 3a), which may be placed in
openings 23 to attach film 12 to the guide rollers. The teeth
preferably maintain the engagement between the film and the guide
rollers as the film is moved through the cassette. Alternately, the
guide rollers may be smooth and a frictional engagement between
guide rollers 15 and film 12 is sufficient to allow the film to be
moved when the guide rollers are rotated. The slide device may
contain an indention 56 shaped to receive container 13a. Indention
56 preferably surrounds conduit 22 to allow container 13a to be
forced within the indention during release of chemical from the
container into conduit 22.
[0060] FIG. 5 depicts a cassette driver 25a for driving cassette 1.
The bottom or base 52 of the cassette may be attached to the upper
surface of the cassette driver. Shaft 18 preferably extends from
the top of cassette driver 25a through an opening in base 52 of
cassette 1. Shaft 18 is preferably a splined shaft. Shaft 18 is
preferably rotated by a motor 26 at a constant speed. Motor 26 may
be a gear drive motor. Shaft 18 preferably drives the motion of
film 12 and piston 16 such that they are synchronized to allow each
holder 13a in film 12 to be struck by piston 16.
[0061] Heat pad 20 is preferably located on the upper surface of
cassette driver 25a. The cassette may contain an opening in base 52
sized to receive the heat pad such that the slide device engages
the heat pad. The temperature of heat pad 20 is preferably
controlled by thermostat 27. The heating contact leads 28a and 28b,
the thermostat contact leads 29a and 29b, and the motor contact
leads 30a and 30b are preferably located at one end of cassette
driver 25a.
[0062] A pressure-sensitive switch 31 is preferably located on top
of cassette driver 25a When cassette 1 is placed on top of cassette
driver 25a, switch 31 is preferably depressed to activate motor 26.
A timing device 32 is preferably used to stop motor 26 after a
predetermined amount of time that is sufficient to allow a selected
amount of chemicals to be applied to the tissue sample.
[0063] FIG. 6 illustrates a multiple cassette driver unit 25b.
Cassette drivers 25a are preferably located within a driver housing
25c. A cassette 1 may be placed on each cassette driver 25a to
stain several tissue samples simultaneously. The cassettes may be
removed from the cassette driver and the slide device and film may
be replaced before the cassette is reinserted on the cassette
driver to stain a new tissue sample.
[0064] Multiple cassette driver unit preferably includes a control
panel 60 for independently operating each of the cassette drivers.
The control panel preferably includes a plurality of controllers 62
corresponding to each of the cassette drivers. Each controller is
preferably electrically coupled to the motor 26 and heat pad 20.
The controller may comprise a timer 32 for stopping motor 26 after
a predetermined amount of running time. The controller may be
further adapted to adjust the operating speed (rpm) of motor 26 to
adjust the speed at which the film is moved and thus the rate that
chemicals are applied to the tissue sample. The controller
preferably includes manual switches to activate and stop motor 26.
The controller may include a computer and a digital display
indicating the temperature of the heat pad and the amount of time
remaining on timer 32. The temperature of the heat pad may be
adjusted manually using controller 62.
[0065] A system of multiple cassette driver units 25b is depicted
in FIG. 7. A plurality of driver housings 25c preferably hold a
plurality of cassette drivers 25a. Tray 25d preferably supports the
multiple cassette driver units 25b, which are arranged
vertically.
[0066] FIG. 8 depicts an alternate embodiment of the tissue
staining system. The portable cassette 42 preferably includes a
positive displacement slide device 31 into which the tissue sample
may be placed. The portable cassette 42 preferably includes a film
32 which carries containers 33 to a solenoid piston 34. Solenoid
piston 34 is operated by battery 35. The solenoid piston 34
preferably forces a chemical from each container 33 into slide
device 31. Guide rollers 36 preferably rotate to move film 32, and
the guide rollers 36 are preferably operated by a twin drive motor
37. A waste tank 38 is preferably located near the end of slide
device 31 for collecting a chemical which exits slide device 31. A
heat pad 39 is preferably located under slide device 31 which may
control the temperature of chemicals within slide device 31. The
presence or absence of contacts along the length of film 32 may
produce closed or open circuits which in turn prevent or allow the
passage of current. The current may activate the heat pad or
actuate the solenoid. Contacts such as contacts 40 and contacts 41
may be placed in multiple rows along film 32 to allow multiple
circuits to be opened or closed simultaneously or independently as
the contacts pass by and come in contact with switches (e.g.,
switch 66) within cassette 42. The switches may be in electrical
communication with the heat pad or the solenoid.
[0067] Turning to FIG. 9, another embodiment of the tissue staining
system is presented. Cassette 1 is preferably shaped to receive
different elements of the staining system. Cassette 1 preferably
includes a cavity 71 and a removable cap 72 which acts as a closure
device for cavity 71. Cap 72 is preferably shaped to snap into a
wall surrounding a top portion of cavity 71. When cap 72 is
removed, plate 8 may be positioned across the bottom of cavity 71.
Cassette 1 may further include a raised protrusion 74 on sides of
cavity 71. Specimen slide 9 may be placed a predetermined distance
above plate 8 such that a sealing element 68 (i.e., a spacer) is
located between the edges of specimen slide 9 and each protrusion
74 to form a head space 10b between plate 8 and specimen slide 9.
Thus, unlike previously depicted embodiments, specimen slide 9 is
not connected to plate 8 while tissue sample 6 is being stained.
The side of specimen slide 9 which contains tissue sample 6
preferably faces head space 10b. Therefore, tissue sample 6 may
protrude downward from slide 9. Another sealing element 70 may be
placed above the outer edges of slide 9, and cap 72 may be snapped
into cassette 1 such that it abutts sealing element 70. Cap 72
preferably locks specimen slide 9 and plate 8 within cavity 71.
Injection port 17 and relief port 68 are preferably formed within
cassette 1 and communicate with head space 10b. Injection port 17
and relief port 68 are preferably located at opposite ends of head
space 10b to allow a chemical to pass into and out of head space
10b. A waste tank 11a may be disposed within cassette 1 underneath
relief port 21 to collect any chemical exiting from head space
10b.
[0068] Film 12 preferably carries a chemically filled container 13a
to a position adjacent to injection port 17. Piston 16 is
preferably aligned with injection port 17. When cassette 1 is
placed onto cassette driver 25a, cassette driver 25a may
automatically start running because starting switch 31 is triggered
by the pressure of cassette 1. Splined shaft 18 preferably extends
up from cassette driver 25a and into cassette 1 where it rotates to
cause cam 59 to displace piston 16. Piston 16 may push container
13a against a sharp end of injection port 17. Members 76 may help
guide piston 16 so that it remains aligned with container 13a. The
pointed end of injection port 17 may puncture the wall of container
13a, forming an opening in the wall. The chemical within container
13a may then flow freely into injection port 17 through this
opening. Heat pad 20 of cassette driver 25a may be used to heat
chemicals passing through head space 10b since it may be positioned
under plate 8. After film 12 has delivered all chemicals necessary
for the staining procedure, a mounting medium may be injected into
head space 10b to form a glue-like substance between specimen slide
9 and plate 8. The "mounting medium" is taken to mean any
translucent substance capable of adhering to slide 9 and to plate
8. Thus, specimen slide 9 and plate 8 are permanently connected to
form a unit which can be removed from cassette 1 and placed under a
microscope for inspection.
[0069] FIG. 10 depicts another embodiment of a tissue staining
system which is similar to the embodiment of FIG. 9, with a few
exceptions. For example, an upper portion 77 of cassette 1 may be
folded back, allowing access to a cavity 71 within cassette 1. One
end of upper portion 77 may be lifted upward (shown by dashed line
78) while the opposite end remains attached to cassette 1 so that
it works like a door. The end of upper portion 77 attached to
cassette 1 may be flexible to enable the lifting mechanism. Upon
opening upper portion 77, a sealing element 80 followed by specimen
slide 9 may be placed across the bottom of cassette 1. Slide 9 is
preferably oriented so that the side containing tissue sample 6
faces upward. Cover plate 9 may be placed a predetermined distance
above plate 9 such that sealing element 68 is located underneath
the edges of plate 9, forming head space 10b between slide 9 and
cover plate 8. Sealing element 70 is then preferably placed on top
of portions of slide 9. Upper portion 77 may be seated upon sealing
element 70 and cover plate 8 when it is closed. The operation of
this system is basically the same as that of the system depicted in
FIG. 9.
[0070] Turning to FIG. 11, an embodiment of a chemical container
13a is depicted. Chemical container 113a is preferably connected to
film 12 and includes a cavity 81. Hereafter, "cavity" is taken to
mean a space and a wall partially enclosing the space. Container
13a may have protrusions 82 spaced along an inner surface of a wall
83 surrounding a portion of cavity 81. Container 13a may be filled
with a chemical and then sealed to prevent the chemical from
escaping. A container cap 86 may be used to seal container 13a. A
portion of cap 86 may be disposed within cavity 81 to plug
container 13a A second set of complementary protrusions 84
preferably line a portion of the outer surface of cap 86.
Protrusions 84 are preferably shaped to fit between protrusions 82
so that cap 86 may be locked to the inner surface of wall 83. A
slit 87 preferably extends through the center of cap 86 and
preferably terminates at another wall 88. When cap 86 is positioned
within cavity 81, a chemical within container 13a may enter slit
87. However, wall 88 preferably prevents the chemical from exiting
from slit 87. During injection of a chemical into cassette 1, wall
88 may be forced against a pointed end of conduit 22, causing wall
88 to be punctured and releasing a chemical into conduit 22.
[0071] Turning to FIG. 12a, a portion of cassette 1 and a portion
of film 12 are illustrated in an embodiment. Cassette 1 may have a
substantially V-shaped edge 94. This edge is preferably sharp
enough to cut through wall 88 which protrudes from a side of each
container 13a. In one embodiment, film 12 carries containers 13a
sequentially to injection port 17. As wall 88 of each container 13a
comes into contact with edge 94, wall 88 may be removed from
container 13a via edge 94 cutting through it Container 13a
preferably abutts surface 92 of cassette 1 to prevent chemicals
within container 13a from escaping through the opening created by
the removal of wall 88. Then, as container 13a passes by injection
port 17, a chemical may flow from container 13a into injection port
17 via the opening. Further, a cavity 90 of container 13a is
preferably located opposite to the opening. A piston may be
propelled into cavity 90 to help force the chemical into injection
port 17. Cavity 90 preferably includes a flexible wall which does
not break open under pressure of the piston. FIG. 12b depicts a
cross-sectional view of the top of film 12 having containers
13a.
[0072] Experiment
[0073] The slide device depicted in FIG. 3 was used to apply
various chemicals to tissue samples. Tissue samples were placed
into eight positive displacement slide devices having a specimen
slide and a cover plate. Each of the slide devices contained a head
space between the specimen slide and the cover plate that had a
width ranging from 0.0005 inch to 0.004 inch. Sixteen control
tissue samples that were each three micrometers thick were mounted
in pairs on the eight specimen slides.
[0074] Each of the tissue samples was stained as follows using
quality-controlled, standardized immunohistochemistry procedures.
The slide device and tissue samples were dried at 58.degree. C. for
one hour. Pure xylene having essentially no dissolved contaminates
of paraffin was injected under pressure into the slide device three
sequential times. The third injection of xylene remained in the
head space of the slide device for ten minutes before a 100%
alcohol composition was injected into the head space. The injection
of the alcohol through the injection port forced the excess xylene
out of the head space. Three minutes later, peroxide/methanol was
injected into the head space to prevent blood and other portions of
the tissue from interacting with chemicals which would subsequently
be injected. The peroxide remained in the head space for thirty
minutes.
[0075] The tissue was then rehydrated via injection of a 95%
alcohol solution into the head space. After three minutes a
deionized water rinse was injected into the head space. Then tissue
revival was performed by heating the slide device and sequentially
injecting an immunohistochemistry (IHC) rinse into the head space
three times. Immediately after injecting the third rinse, a primary
antibody for staining the tissue was injected into the head space.
The primary antibody remained in the head space for fifteen
minutes. An injection of an IHC wash buffer was then performed and
immediately followed by an injection of a secondary antibody known
as link into the head space. After the link had been in the head
space for ten minutes, an IHC wash buffer was injected again.
Immediately thereafter, a chemical known as label was injected into
the head space where it remained for ten minutes.
[0076] Next, an IHC wash buffer was injected into the slide device.
Immediately thereafter, a chromogen was injected into the head
space to colorize the stains. An optimum 4:1 ratio of desired
stained tissue parts to undesired background stained parts was
reached after a time period that ranged from approximately thirty
seconds to five minutes among the tissue samples, and then a
deionized water rinse was injected into the head space. The ratio
of desired stained tissue parts to background stained parts was
determined using a standardized scoring system well known to those
skilled in the art in which 4 is the highest intensity of staining
and 1 is the lowest intensity of background. A counterstain was
injected to produce color contrast. The counterstain remained in
the head space for forty-five seconds and then a deionized water
rinse was injected into the head space. Finally, a translucent
mounting medium was injected into the head space which acted as a
"glue" to seal the cover plate to the head space permanently. The
specimen slide and cover plate could then be placed under a
microscope as a unit.
[0077] The above experiment required only small, efficient amounts
of a chemical for staining tissue. Due to varied head space
distances the required injected volumes varied from 50 microliters
to 200 microliters. The deionized water rinses were quick and
completely cleaned the head spaces because previous chemicals were
positively displaced from the slide devices. Further, uniform and
complete staining was achieved on all sixteen tissue samples.
[0078] The above-mentioned results suggest that
positive-displacement injection of chemicals such as antibodies and
buffers is a viable method for the procedures of
immunohistochemisty. The positive displacement of a previously
injected chemical from a slide device was found to prevent dilution
or mixing between an injected chemical and the previously injected
chemical. Chemical waste is reduced as compared to conventional
methods, and the flow of chemicals under pressure over tissue tends
to cause the tissue to be stained evenly and completely.
[0079] Further modifications and alternative embodiments of various
aspects of the invention will be apparent to those skilled in the
art in view of this description. Accordingly, this description is
to be construed as illustrative only and is for the purpose of
teaching those skilled in the art the general manner of carrying
out the invention. It is to be understood that the forms of the
invention shown and described herein are to be taken as the
presently preferred embodiments. Elements and materials may be
substituted for those illustrated and described herein, parts and
processes may be reversed, and certain features of the invention
may be utilized independently, all as would be apparent to one
skilled in the art after having the benefit of this description of
the invention. Changes may be made in the elements described herein
without departing from the spirit and scope of the invention as
described in the following claims. For example, integrated circuit
devices which embody central processing units and pre-programmed
and/or re-programmable memory may be utilized in the cassette
and/or drive unit to control the function of motors, piston
actuators, heating pads, and status displays. Such electronic
control methodologies could supplement or replace mechanical
methodologies described herein.
* * * * *