Note: Descriptions are shown in the official language in which they were submitted.
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HEAT BLOCKS AND HEATING
RELATED APPLICATIONS
This application claims priority to U.S. Provisional Patent Application Serial
Number
60/691,124, filed on June 16, 2005, which is hereby incorporated by reference
in its entirety.
FIELD
The embodiments disclosed herein relate to heating samples of biological
material,
and more particularly heating and thermal cycling of DNA samples to accomplish
a
polymerase chain reaction, a quantitative polymerase chain reaction, a reverse
transcription-
polymerase chain reaction, an immuno-polymerase chain reaction, or other
nucleic acid
amplification types of experiments.
BACKGROUND
Techniques for thermal cycling of DNA samples are known in the art. By
performing
a polymerase chain reaction, DNA can be amplified. It is desirable to cycle a
specially
constituted liquid biological reaction mixture through a specific duration and
range of
temperatures in order to successfully amplify the DNA in the liquid reaction
mixture.
Thermal cycling is the process of melting DNA, annealing short primers to the
resulting
single strands, and extending those primers to make new copies of double
stranded DNA.
The liquid reaction mixture is repeatedly put through this process of melting
at high
temperatures and annealing and extending at lower temperatures.
In a typical thermal cycler, a biological reaction mixture including DNA will
be
provided in a large number of sample wells on a thermal block assembly. It is
desirable that
the samples of DNA have temperatures throughout the thermal cycling process
that are as
uniform as reasonably possible. Even small variations in the temperature
between one
sample well and another sample well can cause a failure or undesirable outcome
of the
experiment. For instance, in quantitative PCR, one objective is to perform PCR
amplification
as precisely as possible by increasing the amount of DNA that generally
doubles on every
cycle; otherwise there can be an undesirable degree of disparity between the
amount of
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resultant mixtures in the sainple wells. If sufficiently uniform temperatures
are not obtained
by the sample wells, the desired doubling at each cycle may not occur.
Although the
theoretical doubling of DNA rarely occurs in practice, it is desired that the
amplification
occurs as efficiently as possible.
In addition, temperature errors can cause the reactions to improperly occur.
For
example, if the samples are not controlled to have the proper annealing
temperatures, certain
forms of DNA may not extend properly. This can result in the primers in the
mixture
annealing to the wrong DNA or not annealing at all. Moreover, by ensuring that
all samples
are uniformly heated, the dwell times at any temperature can be shortened,
thereby speeding
up the total PCR cycle time. By shortening this dwell time at certain
temperatures, the
lifetime and amplification efficiency of the enzyme are increased. Therefore,
undesirable
temperature errors and variations between the sample well temperatures should
be decreased.
Prior art heat blocks composed of all metal can be expensive. In metal
machined
blocks or metal, thick base, electro-formed blocks, the primary heat transfer
path, from the
heating means to the well cavity, is limited by the wall area of the well.
That is, the heat
must move from the bottoin of the block through the wall area of the well to
heat the well
cavity. To promote heat distribution the well wall is made as thin as possible
to maximize the
heating ramp rate of the well cavity.
In light of the foregoing, there is a need for a thermal cycling apparatus and
method
that enhances temperature uniformity of the sample wells to improve the
efficiency or
accuracy of processing samples. Thus, there is a need in the art for an
apparatus and method
for a non-metal block thermal system for thermal cycling a plurality of
samples.
SUMMARY
In accordance with one aspect of the invention, an apparatus and method are
provided
for a non-metal block thermal system comprising a top plate having a non-metal
material
wherein the top plate comprises a plurality of heating wells, each heating
well sized to
accommodate a plurality of sample tubes containing san-iples. A bottom plate
engages to the
top plate to form a heat block. A plurality of heat transfer pins extend from
the bottom plate
to the top plate. The plurality of heat transfer pins deliver heat to the
plurality of heating
wells. The non-metal material of the top plate may be molded to engage the
heat transfer
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pins and increase heating efficiency. The heat block may be used for efficient
thermal
cycling of biological samples.
One heat block in accordance with this aspect of the invention comprises a top
plate
comprising a non-metal material wherein the top plate comprises a plurality of
heating wells
for accepting a plurality of sample tubes; a bottom plate engaging the top
plate; and a
plurality of heat transfer pins extending from the bottom plate to the top
plate to transfer heat
from the bottom plate to the top plate.
One heat block in accordance with this aspect of the invention comprises a top
plate
comprising a non-metal core with a metal coating on an outer surface and a
bottom plate
comprising a metal.
The present invention also provides a method of thermal cycling samples
comprises
placing a plurality of sample tubes containing samples in a sample retainer,
rotating a rotating
heat assembly to position a top plate of a heat block below the sample
retainer, the top plate
comprising a plurality of heating wells; raising the rotating heat assembly to
bring the
plurality of heating wells of the top plate in thermal contact with the
plurality of sample
tubes; heating the heat block to a first temperature for a first time period
to control a
temperature of the samples in the plurality of sample tubes; and lowering the
rotating heat
assembly so the plurality of heating wells of the top plate are no longer in
thermal contact
with the plurality of sample tubes.
The present invention further provides a method for heating a plurality of
heating
wells comprises providing a heat block having a top plate comprising a non-
metal material
and a bottom plate with a plurality of heat transfer pins engaging a plurality
of heating wells
of the top plate; placing the plurality of heating wells in thermal contact
with a plurality of
sample tubes; heating the bottom plate of the heat block with a heater; and
transferring heat
from the bottom plate to the plurality of heating wells by the plurality of
heat transfer pins.
In accordance with a second aspect of the invention, one or more thermally
conductive materials are used to form at least one plate of a heat block. One
heat block in
accordance with this aspect comprises a thermal plate comprising a thermally
conductive
material, the thermal plate including a major upper surface having a
substantially planar area
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and a plurality of heating wells for accepting a plurality of sample tubes;
and a heating plate
engaging the thermal plate and contacting the major upper surface of the
thermal plate.
The tllermally conductive material can be aluminum.
Additionally, the thermal plate can include a boss having a cavity defined
therein to
simulate a temperature response of a biological sarnple. Alternatively, or in
addition, one
heating well of the heat block can be used to measure a simulated biological
fluid sample
temperature.
One heat block in accordance with this second aspect includes a first plate
having a
substantially planar major upper surface and a plurality of heating wells
defined in the major
upper surface for accepting a plurality of sainple tubes; and a second plate
abutting the major
upper surface of the first plate, the second plate being a heating plate
having a body portion
having a plurality of apertures formed therein, where the body portion can be
substantially
planar in shape; an insulative portion surrounding the plurality of apertures;
and a heating
element carried by the heating plate, arranged between insulative portions
thereof.
The heating element can be a resistive heating element, or a tubular conduit
for
carrying heated fluid, and can be secured to a bottom surface of the second
plate, for
contacting the first plate, embedded in the second plate, or formed within in
the second plate.
The insulative portion can be thermally insulative, or electrically
insulative.
The heat block can also include connecting portions for connecting the heating
element of the second plate to a heat energy source. The heat energy source
can be an
electrical energy source for providing an electrical current to the heating
element, or can be
an energy source that provides heated fluid to the heating element.
Preferably, the first plate comprises a metal material, and can include
copper,
aluminum, brass and combinations thereof. The metal material of the heat block
can also
include a first metal coated with a second metal.
The present invention also provides a method for heating a plurality of
heating wells
comprising; providing a heat block having a thermal plate comprising a
thermally conductive
material, the thermal plate including a major upper surface having a
substantially planar area
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and a plurality of heating wells for accepting a plurality of sample tubes;
and a heating plate
engaging the thermal plate and contacting the major upper surface of the
thermal plate;
placing the plurality of heating wells in tliermal contact with a plurality of
sample tubes;
heating the thermal plate of the heat block with the heating plate; and
transferring heat from
the thermal plate to the plurality of heating wells through conductive heat
transfer.
In one embodiment, the thermal plate of the heat block is heated prior to the
step of
placing the plurality of heating wells in thermal contact with the plurality
of sample tubes.
The present invention also provides a method of thermal cycling samples
comprises
placing a plurality of sample tubes containing samples in a sample retainer,
rotating a rotating
heat assembly to position a top plate of a heat block below the sample
retainer, the top plate
conlprising a plurality of heating wells; raising the rotating heat assembly
to bring the
plurality of heating wells of the top plate in thermal contact with the
plurality of sample
tubes; heating the heat block to a first temperature for a first time period
to control a
temperature of the samples in the plurality of sample tubes; and lowering the
rotating heat
asseinbly so the plurality of heating wells of the top plate are no longer in
thermal contact
with the plurality of sample tubes, and wherein the heat block comprises a
first plate having a
substantially planar major upper surface and a plurality of heating wells
defined in the major
upper surface for accepting a plurality of sample tubes; and a second plate
abutting the major
upper surface of the first plate, the second plate being a heating plate
having a body portion
having a plurality of apertures formed therein; an insulative portion
surrounding the plurality
of apertures; and a heating element carried by the heating plate, arranged
between insulative
portions thereof.
The heat block improves heat transfer, efficiently processes samples, and
allows for
substantial cost savings through the use of a top plate comprising a non-metal
material.
BRIEF DESCRIPTION OF THE DRAWINGS
The presently disclosed embodiments will be further explained with reference
to the
1 attached drawings, wherein like structures are referred to by like numerals
throughout the
several views. The drawings are not necessarily to scale, the emphasis having
instead been
generally placed upon illustrating the principles of the presently disclosed
embodiments.
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FIG. I shows an assembly view of a heat block wherein a top plate of the heat
block
is disengaged from a bottom plate of the heat block.
FIG. 2 shows a perspective view of a bottom plate of a heat block.
FIG. 3 shows a top perspective view of a top plate of a heat block.
FIG. 4 shows a bottom perspective view of a top plate of a heat block.
FIG. 5 shows a cut away view of a heat block wherein a top plate is engaged to
a
bottom plate.
FIG. 6 shows an alternative embodiment of the heat block.
FIG. 7 shows a view of a block heater of a heat block.
FIG. 8 shows a perspective view of a central tube of an embodiment wherein
multiple
heat blocks may engage the central tube.
FIG. 9 shows a perspective view of a rotating heat assembly ivherein a
plurality of
heat blocks engage the central tube.
FIG. 10 is a top perspective view of a thermal plate in accordance with the
invention.
FIG.11 is a bottom perspective view of the thermal plate of FIG. 10.
FIG. 12 is a bottom perspective view of a heating plate in accordance with the
invention.
FIG. 13 is top perspective view of a heat block assembly in accordance with
the
invention, illustrating the thermal plate of FIGS. 10 and 1 I and the heating
plate of FIG. 12 in
an abutting configuration.
FIG. 14 is a perspective view of a rotating heat assembly including three heat
block
asseinblies as shown in FIG. 13.
FIG. 15 shows a perspective view of a rotating heat assembly as part of a
processing
apparatus.
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FIG. 16 shows a close up view of a rotating heat assembly as part of a
processing
apparatus.
FIG. 17 shows a perspective view of a rotating heat assembly as part of a
processing
apparatus.
While the above-identified drawings set forth presently disclosed embodiments,
other
embodiments are also contemplated, as noted in the discussion. This disclosure
presents
illustrative embodiments by way of representation and not limitation. Numerous
other
modifications and embodiments can be devised by those skilled in the art which
fall within
the scope and spirit of the principles of the presently disclosed embodiments.
DETAILED DESCRIPTION
A heat block is capable of delivering a desired amount of heat uniformly and
efficiently to a plurality of samples. The heat block comprises a metal and/or
a non-metal
material to increase the efficiency of processing samples while reducing the
cost of
constructing the heat block.
Therrnal cyclers are the programmable heating blocks that control and maintain
the
temperature of the sample through the three temperature-dependent stages that
constitute a
single cycle of PCR: template denaturation; primer annealing; and primer
extension. These
temperatures are cycled up to forty times or more to obtain amplification of
the DNA target.
Thermal cyclers use different technologies to effect temperature change
including, but not
limited to, peltier heating and cooling, resistance heating, and passive air
or water heating.
Thermal cycling of DNA can accomplish a polymerase chain reaction (PCR), a
quantitative polymerase chain reaction (qPCR), a reverse transcription-
polymerase chain
reaction (RT-PCR), a reverse transcription-quantitative polymerase chain
reaction (RT-
qPCR), immuno-polymerase chain reaction (I-PCR), or other nucleic acid
amplification types
of experiments.
A heat block is shown generally at 67 in FIG. 1. The heat block 67 comprises a
top
plate 83 and a bottom plate 75. In this embodiment, the top plate 83 comprises
a non-metal
material. The top plate 83 may be composed of any plastic material which may
be molded
and has physical properties sufficient to satisfy the structural and
temperature requirements.
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The top plate 83 comprises a high-temperature moldable plastic. The top plate
83 may
comprise polyphenylene sulfide (PPS), polyetherimide (PEI) or other similar
materials
known to those skilled in the art.
In certain embodiments, the top plate 83 comprises a metal coating over a non-
metal
material. The metal coating is on an outer surface of the top plate 83. The
metal coating acts
as a wear surface and permits easy cleaning of the top plate 83. The metal
coating also
promotes the sensitivity of the instrument, when used to obtain quantitative
data, by
providing a more optically reflective surface as compared to the molded
plastic surface. The
metal coating or plating may comprise copper, nickel, chromium, gold, or a
combination of
nzultiple metals or other metals known to those skilled in the art. The metal
coating may be
applied using coating methods known in the art including, but not limited to,
bath plating,
physical, chemical, or ion vapor deposition, or other coating methods known in
the art.
The bottom plate 75 coinprises a conductive material, preferably a metal. The
bottom
plate 75 may be prepared by casting, machining, forging, metal injection
molding or other
methods known in the art. In an embodiment, the bottom plate 75 comprises
aluminum. The
bottom plate 75 may comprise copper, silver, aluminum alloy, other castable
alloys or other
similar materials known to those skilled in the art. The bottom plate 75 may
comprise a
plurality of metals. Those skilled in the art will recognize that the bottom
plate may comprise
a variety of conductive materials and be within the spirit and scope of the
presently disclosed
embodiments.
The design of the top plate 83 in relation to the bottom plate 75 allows for
heat to be
more evenly distributed throughout a sample which leads to more uniform,
efficient and
reliable results as compared to those obtained through the use of prior art
heat blocks. The
use of a non-metal material for the top plate 83 is cost effective. Processing
a large number
of samples often requires the use of a large number of heat blocks which
increases expenses.
The presently disclosed embodiments allow for substantial cost savings by
using heat blocks
that can have a non-metal top plate. As such, the presently disclosed
embodiments provide
both cost savings and improved results.
FIG. 1 shows an assembly view of a heat block wherein the top plate 83 of the
heat
block 67 is disengaged from the bottom plate 75 of the heat block 67. A
plurality of heat
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transfer pins 77 extend from the bottom plate 75. The plurality of heat
transfer pins 77 may
be formed witli the bottom plate 75. Alternatively, the plurality of heat
transfer pins 77 may
be added to the separate bottom plate 75. The plurality of heat transfer pins
77 may be
composed from a different material than the bottom plate 75. The plurality of
heat transfer
pins 77 may be different shapes including, but not limited to, square,
rectangular, circular,
oval, and other shapes. The plurality of heat transfer pins 77 may be etched,
roughened,
grooved, notched or formed with any other surface effect such that the surface
area for each
pin is increased. This may be useful to promote a mechanical connection with a
plurality of
ribs 87 of the top plate 83.
The plurality of heat transfer pins 77 deliver heat to a plurality of heating
wells 85.
The plurality of heat transfer pins 77 distribute heat evenly to each
individual sample. The
plurality of heat transfer pins 77 transfer heat to the sides of the plurality
of heating wells 85
that contain the plurality of sample tubes 39. The plurality of heat transfer
pins 77 act as an
interface to deliver heat along the entire length of the sample tube 39.
Delivering heat to the
sides of the heating well 85 distributes heat in a saniple better than
delivering heat to the
sample exclusively through the bottom of the heating well 85. The heating well
85 is molded
so that the heating well 85 better engages the heat transfer pins 77 and
therefore allows for
heat to be efficiently delivered to the heating well 85 from the sides of the
heating well 85
through thermal conduction. The flexibility and moldability of the non-metal
material of the
top plate 83 to engage the heat transfer pins 77 provides uniform heating to
the sample and
increases processing efficiency. The heating well 85 may have the plurality of
ribs 87 to
engage the plurality of heat transfer pins 77 and promote uniform heat
distribution.
The heating ramp rate is important as a user feature since it impacts the
speed at
which a user can conduct a biological experiment. As the heating well wall
thickness is
reduced, to reduce the amount of heated mass, the area for the heat path is
reduced as well.
As the heating well wall thickness is reduced, the temperature gradient within
a single
heating well cavity increases. This single well temperature gradient may
become as great or
greater than the temperature gradient across the entire sample block for thin
well walls. The
plurality of heat transfer pins 77 combined with the top plate 83 allows the
heat transfer path
to enter the heating well 85 from the sides of the heating wel185. The heat
travels from the
plurality of heat transfer pins 77 through the plurality of ribs 87 which
engage the heating
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well 85 on the underside of the top plate 83. In an embodiment, the heating
well 85 has four
ribs 87. More or less ribs, or ribs with other shapes and orientations may be
used to transfer
the heat to the heating well in alternative embodiments, given fabrication and
cost
considerations. The presently disclosed embodiments provide a significant
increase in the
heat transfer area from the heating means to the heating well.
The geometry of the plurality of ribs 87 contained in the top plate 83 and the
plurality
of heat transfer pins 77 in the bottom plate 75 inay be optimized to promote
both single well
temperature uniformity and also complete heat block temperature uniformity.
The
optimization is obtained by the size and orientation of the rib draft angle,
rib thickness versus
heating well position; pin size, pin draft angle, and other design dimensions
and
characteristics.
The amount of pin 77 to rib 87 contact area versus well position forms a
design
configuration for excellent temperature unifornlity across the heat block. In
an embodiment,
for internal wells, four pins 77 contact each heating well rib 87 and the four
pins 77 share
contact with two heating well ribs 87 each. In an embodiment, for edge wells,
four pins 77
contact each heating well rib 87, but only three pins 77 share contact with
two heating well
ribs 87 each. The fourth pin 77 does not share contact with another well and
therefore more
heat from the fourth pin 77 is available for the edge heating wells 85 to help
counteract the
inherently cooler edge temperature of a heated rectangular body. The corner
heating wells 85
benefit from four pins 77 which contact each heating well rib 87. Only two of
the pins 77
share contact with another heating well. The other two pins 77 do not share
contact with
another well and even more heat is available for the corner wells to
counteract the inherently
cooler corner temperature of a heated rectangular body. Those skilled in the
art will
recognize the number of pins and ribs may vary and still be within the spirit
and scope of the
presently disclosed embodiments.
As shown in FIG. 1, the bottom plate 75 comprises a plurality of recesses 78.
The
plurality of recesses 78 have varying depths and are symmetrical about the
bottom plate.
Each recess 78 is designed to promote the temperature uniformity across the
heat block when
engaged with the top plate 83. The plurality of recesses 78 form an efficient
way to promote
temperature uniformity since they are efficiently formed via machining,
casting, or other
fabrication methods known in the art. Other shapes and patterns of recesses,
or no recesses,
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may be used in alternative embodiments. FIG. 2 shows a perspective view of the
bottom
plate 75 of the heat block 67. FIG. 3 shows a top perspective view of a top
plate of a heat
block 67. FIG. 4 shows a bottom perspective view of the top plate 83 of the
heat block 67.
As shown in FIG. 5, the top plate 83 engages the bottom plate 75 to form the
heat
block 67. The top plate 83 may be connected to the bottom plate 75 by any
mechanical
engagement known in the art including, but not limited to glue, welding, snap
fit, shrink fit,
press fit, epoxy, adhesives and other mechanical fasteners known in the art
and be within the
spirit and scope of the presently disclosed embodiments. An alternative
configuration could
utilize a process where the bottom plate is used as a mold insert. In this
way, the top plate
could be molded and attached to the bottom plate during the molding process.
In addition to the top plate of the heat block having ninety-six heating wells
shown in
FIGS. 1-5, the heat block may have one or more dedicated wells 97 for
measuring a
simulated biological fluid sample temperature. The heat block may have one,
two, three, four
or more dedicated wells 97 for measuring a simulated biological fluid sample
temperature. In
an embodiment, the dedicated wells 97 for measuring a simulated biological
fluid sample
temperature are located along the edges of the heat block 67. Those skilled in
the art will
recognize that the dedicated wells 97 for measuring a simulated biological
fluid sample
temperature could be located along the edges, in the middle or at other
locations in the top
plate 83 of the heat block 67. A control system where a simulated biological
fluid
temperature measurement is used to make accurate transitions between
biological sample
fluid temperatures may also be used.
Although FIGS. 1-5 show the heat block 67 with a 96 well configuration, those
skilled
in the art will recognize that 48 well, 384 well, 1536 well, and other
multiple well heat blocks
are within the spirit and scope of the presently disclosed embodiments.
FIG. 6 shows an alternative embodiment of the heat block 67 for use with 384
well
plates. The top plate 83, which can be made of a non-metal material may be
molded in a
variety of forms to best match commercially available sample container sizes
and
configurations. The 384 well format is often used for higher throughput of DNA
samples.
The bottom plate 75 may be cast or otherwise formed to support a variety of
configurations.
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FIG. 7 shows a view of a block heater 107 for heating the heat block 67. The
heater
107 is positioned below the bottom plate 75 of the heat block 67. Heat is
generated by the
heater 107 and transferred to the bottom plate 75. The heat transfer pins 77
of the bottom
plate 75 then transfer the heat to the heating wells 85 in the top plate 83
that hold the sample
tubes containing samples. Placing the heater 107 below the bottom plate 75
provides an
efficient and uniform transfer of heat to the samples. The heater 107 may heat
the samples by
resistance heating, peltier heating and cooling passive air or water heating
and other heating
and cooling methods known in the art and be within the spirit and scope of the
presently
disclosed embodiments.
The heat block 67 may be used in a variety of ways. The heat block 67 may be
used
; in isolation to process a plurality of samples. Alternatively, the heat
block 67 may be used
with a plurality of additional heat blocks 67 in order to perform various
processes (i.e., a first
heat block delivers heat to the samples for a first time period, a second heat
block delivers
heat to the samples for a second time period, a third heat block delivers heat
to the samples
for a third time period, etc.).
The following discussion illustrates one use of the heat block 67. The
following
discussion is in no way meant to limit the use of the heat block of the
presently disclosed
embodiments. Those skilled in the art will recognize that various uses of the
heat block are
within the spirit and scope of the presently disclosed embodiments.
A plurality of heat blocks 67 may be used in conjunction to process a
plurality of
samples. The plurality of heat blocks may be utilized in a single process
wherein each heat
block may be operated at a different temperature. The plurality of heat blocks
may be
engaged to a central tube thereby creating a rotating heat assembly. The
rotating heat
assembly may be engaged to a processing apparatus. The rotating heat assembly
may be
rotated so that any of the heat blocks engaged to the rotating heat assembly
may be positioned
parallel to a plurality of sample tubes containing samples. The rotating heat
assembly may be
moved up and down in order to bring either heat block into thermal contact
with the plurality
of sample tubes. The presently disclosed embodiments include a method of
improving PCR
efficiency by using the apparatus of the presently disclosed embodiments to
rapidly bring a
plurality of heat blocks into and out of thermal contact with the plurality of
sample tubes and
avoiding the problems of raising and lowering the temperature of a single heat
source.
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FIG. 8 shows a perspective view of an embodiment wherein multiple heat blocks
may
engage a central tube 103. The central tube 103 coinprises a cylindrical
geometry. A first
heat block, a second heat block, and a third heat block may engage the central
tube 103.
Those skilled in the art will recognize that the central tube 103 may have any
of a variety of
shapes and geometries and be within the spirit and scope of the presently
disclosed
embodiments.
FIG. 9 shows a perspective view of a rotating heat assembly 65 in whicli the
plurality
of heat blocks engage the central tube 103. In FIG. 9, a first heat block 67,
a second heat
block 69, and a third heat block 71 engage the central tube 103 to produce the
rotating heat
block assembly 65. The rotating heat block assembly 65 may comprise two,
three, four or
more heat blocks and be within the spirit and scope of the presently disclosed
einbodiinents.
Those skilled in the art will recognize that that various methods of engaging
the heat blocks
67, 69, 71 to central tube 103 may be within the spirit and scope of the
presently disclosed
embodiments.
The rotating heat block assembly 65 may engage to a processing apparatus 100.
The
use of the rotating heat block assembly 65, comprising the plurality of heat
blocks 67, 69, 71,
allows for a plurality of samples to be processed rapidly and efficiently.
Those skilled in the
art will recognize that various types of processing assemblies are within the
spirit and scope
of the presently disclosed embodiments.
Reference will now be made to the embodiment illustrated in Figures 10-14,
unless
otherwise noted. Figures 10-14 illustrate an alternate embodiment of a first
plate 200, a
second heat block plate, which is a heating plate 300 and a complete heat
block assembly
400, in accordance with the invention. The first plate 200 can be configured
to be made of a
thermally conductive material, can include a major upper surface having a
substantially
planar area and a plurality of well cavities or "heating wells" 285 for
accepting a plurality of
sample tubes defined therein. The term major upper surface is intended to mean
an upper
surface which constitutes a portion of an upper surface of the first plate
200. For example,
since wells 285 are formed in the first plate 200, and interrupt the otherwise
substantially
planar upper surface, the space between the wells 285 is referred to as a
major upper surface
283. The purpose of the first plate is consistent with that of the top plate,
such as top plate 83
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of Figure 1, described above. However, as is apparent, the configuration
thereof is somewhat
different.
The first plate 200, in accordance witli this embodiment is preferably made of
a heat-
conducting material such as a metal. However, metal-coated materials can be
utilized,
including, but not limited to metal-coated polymers, or metal-plated metals.
Metals such as
copper, nickel, chromium, gold, other suitable materials and combinations
thereof can be
used. In certain embodiments, the first plate 200 comprises a metal coating
over a metal
material. The metal coating is on an outer surface of the first plate 200. The
metal coating
acts as a wear surface and permits easy cleaning of the first plate 200. The
metal coating also
promotes the sensitivity of the instrument, when used to obtain quantitative
data, by
providing a more optically reflective surface as compared to the first plate
surface (which
may also be optically reflective). The metal coating or plating may coinprise
copper, nickel,
chromium, gold, or a combination of multiple metals or other metals known to
those skilled
in the art. The metal coating may be applied using coating methods known in
the art
including, but not limited to, bath plating, physical, chemical, or ion vapor
deposition, or
other coating methods known in the art.
While shown in the accompanying figures as having 96 heating wells 285, first
plate
200 may include 48 well, 384 well, 1536 well, and other multiple well
configurations. In
addition, while shown as being substantially circular in cross section, the
heating wells 285
may be of any shape suitable to the use of first plate 200 in thermal cycling.
The second plate 300 is configured to serve as a heating plate, and is
configured to
abut the major upper surface 283 of the first plate 200, and/or to engage the
first plate. The
second plat preferably includes a plurality of apertures 310, defined therein,
which
correspond to respective wells 285 of the first plate 200. The second plate
can further include
a heating element 320, carried by the plate 300. Optionally, insulative
portions 330 are
provided between the heating element 320 and the edge of each aperture 310.
The purpose of
such insulation is twofold. Firstly, the insulation serves to distribute the
heat from the ,
heating element 320, allowing the edge of the aperture 310, and anything in
contact therewith
to change temperature more gradually than the heating element 320 itself.
Further, since the
insulation is slower to transfer heat, the insulation moderates any
temperature fluctuations of
the heating element 320, or the second plate 300 in general. When the heating
element 320 is
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a resistive electrical heating element, the insulation also helps electrically
insulate the edge of
the apertures 310, through which test tubes will normally pass, from the
electrical current
running through the heating element 320.
This embodiment of first and second plates 200, 300, and the combined heat
block
assembly 400 differ from the foregoing embodiments, and typical heat blocks,
in that the
second plate 300, which is the heating plate, is in contact with an upper
surface of the first
plate. This provides certain advantages over typical heat blocks.
Specifically, since the well
walls 295 of heating wells 285 taper toward the bottom end 297 of each well,
the cross-
section of each wel1285 is greater near the top of the we11285, which is in
direct thermal
communication with the major upper surface 283 of the first plate 200.
Therefore, the upper
portion of the well walls can conduct more heat than the bottom portion of the
well walls. As
configured in this embodiment, since heat will be applied via the second
plate, which is in
contact with the major upper surface 283 of the first plate 200, heat will
more effectively
flow to the far end of the heating we11285, which is the bottom end 297
thereof, than if heat
were to flow in the opposite direction, that is, from the bottom 297 of the
heating well to the
top thereof, near the major upper surface 283. As embodied, improved intra-
well thermal
uniformity is achieved.
A further advantage of this embodiment over certain typical heating blocks, is
that a
negative draft angle in forming for the first plate is not needed. Some
typical thermal plates,
which serve a similar purpose to that of the present first plate 200, include
a heating well
bottom wall that is wider than the top, in order to better contact a heating
element arranged on
the bottom surface thereof. The present invention obviates such negative draft
angle, and the
cumbersome manufacturing processes needed to manufacture such components.
Still a fiuther advantage of the embodiment of Figures 10-14 is that in use,
convective
heat losses are reduced. Since many typical heating blocks provide a heating
element
arranged on a bottom of the heating block, heat can be lost through convection
with the
surrounding environment. The second plate 300 of the present heat block 400,
in use, will be
situated between the first tray 200, which is intentionally heated, and a
cover of a processing
apparatus, such as processing apparatus 100 in Figure 16. Accordingly, little
or no portion of
the second plate 300 is exposed to the surrounding environment or subject to
substantial
convective heat loss. As a result, the temperature uniformity among wells 285
in the first
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plate 200 and the heating block 400 as a whole, is improved, as compared with
a heating
block having an exposed heating element.
Two boss extensions 287, which extend away from the major upper surface 283,
along the long edge 282 of the heat block, are used for heat block retention
in a supporting
frame. These bosses 287 interface with cutouts 455 in a heat block retainer
450 (Figure 14),
to capture and partially locate the heat block 400 in a heat block assembly
500. Along one or
both short edges 284 of the first plate, one or more bosses 289, which can
include a cavity
288 define therein, can be used to interface with a temperature sensor for
simulating the
temperature response of a biological sample mixture in contact with the
heating wells 285,
albeit if througli a vial or test tube wall. While the embodiment of Figures
10-14 includes
two such bosses 289 and cavities 288, fewer than or more than two may be
provided.
Moreover, the precise location of these cavities can be altered given other
system design
considerations, and such temperature sensors can alternatively be provided in
one or more of
the wells 285 of the first tray 200, as described above in connection with
Figures 1-5.
Figure 11 illustrates an underside of the first tray 200, in accordance with
this
embodiment of the invention. As can be seen, substantially cylindrical
recesses 293 are
provided in the bottom surface 291 of the first plate 200, opposite of the
major upper surface
283. These recesses 293 promote heat block temperature uniformity in the
horizontal plane
by distributing the heat block mass in the horizontal plane. Such distribution
can be modified
by selecting appropriate diameter, location, depth or other aspects, or other
attributes to the
recess 293, including use of other shapes, such as polygonal shapes such as
square, hexagonal
and the like, or a hyperbolic shape, such as region 321 of the heating element
of the second
plate 300, as will be described hereinbelow.
The second plate 300, which acts as a heating plate for the subject heating
block 400,
is best illustrated in Figure 12. This heater contains apertures 310 defined
therein, which
correspond to the heating wells 285 provided in the first plate 200. As such,
in use, a sample
tube, such as a vial or test tube can pass through one of the apertures 310,
and into a well 285.
Insulation 330 is provided around the apertures 310, and a heating element 320
passes
between rows of apertures 310 and insulation 330 in a substantially serpentine
manner. As
can be seen, the heating element 320 begins at each end at a connecting
portion 340a, 340b.
from there, the heating element follows a path that passes between each row of
apertures 310,
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until reaching the opposite end. This configuration helps promote temperature
uniformity
throughout the second plate 300, and the heating block 400 as a whole.
Insulating materials that can be used in conjunction with this second plate
300
include, but are not limited to silicone rubber, polyimide (PI), mica,
polyester, nomex, and
other similar materials. Such materials are described in U.S. patent No.
6,878,905, which is
hereby incorporated by reference in its entirety. The insulating materials can
be electrically
insulative, thermally insulative or both electrically and thermally
insulative.
The heating element 320 can include an electric resistive heating material, or
can be
another suitable type of heating element. A warm side of a Peltier junction
can be configured
to be in contact with the major upper surface 283 of the first tray 200.
Alternatively, a fluid-
carrying conduit can be provided, to interface with an external source for
heated or cooled
fluid, such as a heat pump or a hot water supply.
The heating element 320, as illustrated, includes expanded-width regions at
points
located between four apertures 310, such regions having a substantially
hyperbola-shaped
border. Accordingly, heat is provided more evenly to regions near the
circumference of the
apertures 310 and heating wells 285. Connection portions 340a, 340b are
provided to enable
electrical connection of the heating element 320 of the second plate 300 to an
electrical
source.
If desired, the heating element can be substituted for an element that can
provide heat
or remove heat, or alternatively still, only remove heat, depending on the
desired capability of
the heating block. For example, a cool side of a Peltier junction can be used
in place of a
resistive heating element 320. Alternatively still, if a tubular element is
provided to conduct
conditioning fluid, that is, heating or cooling fluid, then a cold fluid, such
as chilled liquid or
refrigerant can be used. If a heat pump system is utilized, then a user need
only select the
desired temperature, and the system will heat or cool the heating block as
necessary.
The heating element 320 and/or cooling element, if so embodied, can be applied
to a
surface of the second plate 300, or can be embedded therein. For example,
fluid-carrying
tubes can be provided on or in the second plate 300. The heating/cooling
elements 320 and
insulation 330 can both be carried by a substrate, or can be mutually joined
without a
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substrate, such that the second plate 300 is consistent in composition
throughout cross-
sections taken parallel to the substantially planar top and bottom surfaces
thereof.
The first plate 200 and the second plate 300 can be joined in any suitable
manner,
such as those set forth above. That is, the first plate 200 can be connected
to the second plate
300 by any mechanical engagement known in the art including, but not limited
to glue,
welding, snap fit, shrink fit, press fit, epoxy, adliesives and other
mechanical fasteners known
in the art and be within the spirit and scope of the presently disclosed
embodiinents. An
alternative configuration could utilize a process where the plates are
integrally formed, for
example, in casting or molding.
Figure 14 illustrates a perspective view of a rotating heat block assembly
500, in
which the plurality of heat blocks 400 engage a central tube 103, similarly to
the embodiment
of Figure 9. The rotating heat block assembly 500 can include two, three, four
or more heat
blocks and be within the spirit and scope of the presently disclosed
einbodiments. Those
skilled in the art will recognize that that various niethods of engaging the
heat blocks 410a,
410b, 410c to central tube 103 may be within the spirit and scope of the
presently disclosed
embodiments.
The rotating heat block assembly 500 can engage to a processing apparatus 100
(See Figure
15, for example). The use of the rotating heat block assembly 500 comprising
the plurality of
heat blocks 410a-410c allows for a plurality of saniples to be processed
rapidly and
efficiently. Those skilled in the art will recognize that various types of
processing assemblies
are within the spirit and scope of the presently disclosed embodiments. FIG.
15, FIG. 16
and FIG. 17 show various views of an embodiment of the processing apparatus
100. FIG. 15
shows a perspective view of the rotating heat assembly 65 as part of the
processing apparatus
100. In FIG. 15, a tube cover 13 is in an open position. The tube cover 13
comprises a tube
cover handle 11 for moving the tube cover 13 from an open position to a closed
position.
The tube cover 13 may include a tube cover heater 15. A sample retainer 43
contains a
plurality of sample wells 38 for receiving a plurality of sample tubes 39. The
plurality of
sample tubes 39 are placed in the plurality of sample wells 38. The sample
wells 38 provide
a means to accommodate a variety of sample tube 39 formats commonly used for
biological
experiments. Some of these sample tube 39 formats include, but are not limited
to, strips of
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eight tubes, flange connected plates of 96 tubes, single tubes, and other tube
configurations
known in the art. A sample carrier 45 supports the sample retainer 43.
FIG. 15 shows the rotating heat assembly 65 engaged with the processing
apparatus
100. The rotating heat assembly 65 comprises a plurality of heat blocks. In
FIG. 15, the
rotating heat assembly 65 comprises three heat blocks: a first heat block 67,
a second heat
block 69 and a third heat block 71.
The first heat block 67 is capable of reaching and maintaining a first
temperature, the
second heat block 69 is capable of reaching and maintaining a second
temperature, and the
third heat block 71 is capable of reaching and maintaining a third
tenlperature. The first
temperature, the second temperature and the third temperature can be the same
or distinct
from one another. In some embodiments, only the first temperature and the
second
temperature are distinct, and the third temperature is the same as the first
temperature or the
second temperature. Those skilled in the art will recognize that each heat
block may reach
and maintain any temperature and be within the spirit and scope of the
presently disclosed
embodiments.
As shown in FIG. 15, the plurality of heat blocks 67, 69, 71 are engaged to
the central
tube 103 wherein the central tube 103 runs from a first slide 37 to a second
slide 53 and the
central axis of the central tube 103 is substantially horizontal.
The plurality of heat blocks 67, 69, 71 each have the top plate 83 with a
plurality of
heating wells 85. While the figures show the heat blocks 67, 69, 71 with the
top plate 83
having 96 heating wells 85, those skilled in the art will recognize that the
heat block may
have 48 wells, 384 wells, 1536 wells, and other numbers of wells and be within
the spirit and
scope of the presently disclosed embodiments. The number of heating wells 85
in the top
plate 83 corresponds to the number of sample wells 38 in the sample retainer
43.
As shown in FIG. 15, the central tube 103 is rotated by a rotation motor 49
that is
mechanically connected to the central tube 103. The central tube 103 is
rotated so the top
plate 83 of the heat block 67, 69 or 71 is below, aligned with and
substantially parallel to the
sample retainer 43. Each sample wel138 of the sample retainer 43 is positioned
above the
heating well 85 of the top plate 83 of the heat block 67, 69 or 71. As will be
discussed in
greater detail below, the rotating heat assembly 65 maybe raised and lowered,
allowing the
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plurality of sample tubes 39 supported in the plurality of sample wells 38 to
be received by
respective ones of the plurality of heating wells 85. The plurality of sample
tubes 39 are thus
in thennal contact with the plurality of heating wells 85 of the top plate 83.
The rotating heat assembly 65 may be raised and lowered to any desired
vertical
position. The rotating heat assembly 65 may be raised and/or lowered manually
or
automatically using a motor 25. Those skilled in the art will recognize that
various
mechanisms and/or motors may be utilized to raise and lower the heat block 67
of the rotating
heat assembly 65 and be within the spirit and scope of the presently disclosed
embodiments.
FIG. 16 shows a close up view of the rotating heat assembly 65 as part of the
processing apparatus 100. FIG. 16 shows a view from below the sample retainer
43. The
rotating heat asseinbly 65 is engaged to the processing apparatus 100. The top
plate 83 of the
second heat block 69 is below, aligned with and substantially parallel to the
sample retainer
43. The plurality of heating wells 85 of the second heat block 69 are
positioned beneath the
plurality of sample tubes 39. The plurality of sample tubes 39 are located
within the plurality
of sample wells 38 of the sample retainer 43. A sample fluid sensor 57 or a
plurality of
sample fluid sensors 57 are operatively connected to the sample retainer 43.
The rotating heat assembly 65 may be rotated to a desired position. As shown
in FIG.
16, a rotational position sensor 63 is in communication with the rotating heat
asselnbly 65.
The rotational position sensor 63 controls and/or indicates the current
rotational position of
the rotating heat assembly 65. The rotating heat assembly 65 may be raised or
lowered to a
desired vertical position. A vertical position sensor 59 is in communication
with the top plate
83 of the heat block 67. The vertical position sensor 59 controls and/or
indicates the current
vertical position of the rotating heat assembly 65. The position sensors 59,
63 provide a
repeatable position signal. The position sensors 59, 63 are selected to
support a position
resolution sufficient for reliable motion. In an embodiment, the position
sensors 59, 63
comprise multiple terminals for wire harn.ess connection. Those skilled in the
art will
recognize that various rotational and/or vertical position sensors known in
the art may be
within the spirit and scope of the presently disclosed embodiments.
FIG. 17 shows a perspective view of the rotating heat assembly 65 as part of
the
processing apparatus 100. In FIG. 17, the tube cover 13 has been lowered to a
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position to cover the sample retainer 43 and the plurality of sample tubes 39.
The tube cover
13 comprises the tube cover heater 15 (shown in FIG. 15). Closing the tube
cover 13 and
activating the tube cover heater 15 heats the plurality of sample tubes 39.
The tube cover
heater 15 provides heat to the plurality of sample tubes 39 to provide a
desired temperature
profile. The tube cover heater 15 may provide a constant temperature or a
variable
temperature during processing of the plurality of sample tubes 39. The tube
cover heater 15
may be used in conjunction with the rotating heat assembly 65 to achieve a
desired
temperature profile of the plurality of sample tubes 39. Those skilled in the
art will recognize
that various temperature profiles are within the spirit and scope of the
presently disclosed
embodiments.
The tube cover 13 comprises a temperature sensor to sense a cover temperature
so that
this temperature may be actively controlled. The temperature sensor may be a
thermistor
engaged to the tube cover 13. The thermistor has multiple lead wires that exit
the tube cover
13 and are operatively connected to the thermal system.
A temperature sensor may be positioned in a sainple container within a sample
well
38 of the sample retainer 43 to measure the temperature of the heat block 67.
The
temperature data from the temperature sensor is sent to a controller which
will then adjust the
amount of heat provided by the heat source. The temperature sensor may be a
thermistor.
The thermistor accurately controls the components involved in a temperature
transition.
Those skilled in the art will recognize that thermocouples, resistance
temperature detectors
(RTD) or other temperature sensors known in the art are within the spirit and
scope of the
presently disclosed embodiments.
The presently disclosed embodiments provides a method of performing thermal
cycling comprising placing at least one sample tube 39 in at least one sample
well 38 of the
sample retainer 43 engaged to a main frame of the processing asseinbly 100.
The rotating
heat assembly 65 is rotated so the top plate 83 of the first heat block 67 is
positioned below
the sample retainer 43. The top plate 83 comprises at least one heating
wel185. The rotating
heat assembly 65 is raised to bring the heating well 85 into thermal contact
with the plurality
of sample tubes 39 and allow the heating wells 85 remain in thermal contact
with the plurality
of sample tubes 39 for a first time period at a first temperature. The
rotating heat assembly
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65 is lowered to separate the plurality of heating wells 85 from the plurality
of sample tubes
39 so the heating wells 85 and sample tubes 39 are no longer in thermal
contact.
The second heat block 69 is heated to a second temperature and the second heat
block
69 is rotated into a position below the sample retainer 43. The rotating heat
assembly 65 is
raised to bring the second heat block 69 into thermal contact with the
plurality of sample
tubes 39. The plurality of heating wells 85 remain in thermal contact with the
plurality of
sample tubes 39 for a second time period at a second temperature so the
samples in the
plurality of sample tubes 39 attain a desired temperature profile. The
rotating heat assembly
65 is lowered after the second heat block 69 has heated the plurality of
sample tubes 39 for a
sufficient time period. The rotating heat assembly 65 is lowered to separate
the plurality of
heating wells 85 from the plurality of sample tubes 39 so the heating wells 85
and sample
tubes 39 are no longer in thermal contact.
The third heat block 71 is heated to a third temperature and the third heat
block 71 is
rotated into a position below the sample retainer 43. The rotating heat
assembly 65 is raised
to bring the third heat block 71 into thermal contact with the plurality of
sample tubes 39.
The plurality of heating wells 85 remain in thermal contact with the plurality
of sample tubes
39 for a third time period at a third temperature so the samples in the
plurality of sample
tubes 39 attain a desired temperature profile. The rotating heat assembly 65
is lowered after
the third heat block 71 has heated the plurality of sample tubes 39 for a
sufficient time period.
The rotating heat assembly 65 is lowered to separate the plurality of heating
wells 85 from
the plurality of sample tubes 39 so the heating wells 85 and sample tubes 39
are no longer in
thermal contact.
The method maybe repeated for multiple heat blocks. The heat blocks may
operate at
a constant temperature or a variable temperature while in thermal
communication with the
plurality of sample tubes. The heat blocks of the rotating heat assembly may
operate at
varying temperature and for varying time periods for the samples to reach a
desired
temperature profile.
Other sample holding structures such as slides, partitions, beads, channels,
reaction
chambers, vessels, surfaces, or any other suitable device for holding a sample
can be used
with the presently disclosed embodiments. The samples to be placed in the
sample holding
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structure are not limited to biological reaction mixtures. Samples could
include any type of
product for which it is desired to heat and/or cool, such as cells, tissues,
microorganisms or
non-biological product.
Each sample tube 39 can have a corresponding cap for maintaining the
biological
reaction mixture in the sample tube. The caps are typically inserted inside
the top cylindrical
surface of the sample tube. The caps are relatively clear so that light can be
transmitted
through the cap. Similar to the sample tubes, the caps are typically made of
molded
polypropylene, however, other suitable materials are acceptable. Each cap has
a thin, flat,
plastic optical window on the top surface of the cap. The optical window in
each cap allows
radiation such as excitation light to be transmitted to the DNA samples and
emitted
fluorescent light from the DNA to be transmitted back to an optical detection
system during
cycling.
Heat blocks in accordance with the invention can be used with thermal cyclers
of
various inakes and models, and is not limited to use in a thermal cycler as
exemplified in
FIGS. 8, 9 and 14-17. Other thermal cycler systems and methods of detecting
the
fluorescence from a PCR reaction could also benefit from a heat block of the
presently
disclosed embodiments. For example, the heat block could be used with the
apparatus for
thermally cycling samples of biological material described in assignee's U.S.
Patent No.
6,657,169, and the entirety of this patent is hereby incorporated herein by
reference. The heat
block can also be used with the Mx3000P Real-Time PCR System and the Mx4000
Multiplex
Quantitative PCR System (commercially available from Stratagene California in
La Jolla,
CA) using a tungsten halogen bulb that sequentially probes each sample,
detected with a
photomultiplier tube. In addition, the heat block could be used with thermal
cyclers
incorporating any or all of the following: a tungsten halogen bulb that
sequentially probes
each sample; a scanning optical module; stationary light emitting diodes
(LEDs) for each
well and the same detector for all wells; stationary samples, light sources,
and detectors;
stationary LEDs and a detector to probe spinning samples sequentially; a
tungsten halogen
bulb to illuminate the entire plate and a charge-coupled device detection of
the entire plate; a
stationary light source and multiple detectors sampling spinning capillaries
sequentially; a
stationary laser and detector that sequentially probes stationary samples
using independent
fiber optics collecting light from each sample; a tungsten halogen bulb to
illuminate the entire
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plate and charge-coupled device detection of the entire plate, and other
thermal cyclers
known in the art.
All patents, patent applications, and published references cited herein are
hereby
incorporated by reference in their entirety. It will be appreciated that
various of the above-
disclosed and other features and functions, or alternatives thereof, may be
desirably combined
into many other different systems or applications. Various presently
unforeseen or
unanticipated alternatives, modifications, variations, or improvements
tllerein may be
subsequently made by those skilled in the art which are also intended to be
encompassed by
the following claims.
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