Thermal cycler for automatic performance of the polymerase chain reaction with close temperature control
First Claim
1. An apparatus for controlled automated performance of polymerase chain reactions in at least one sample tube containing a known volume of a liquid sample mixture, which apparatus comprises:
- a. A sample block having at least one well for said at least one sample tube,b. a computing apparatus,c. heating and cooling means controlled by said computing apparatus for changing the temperature of said sample block, andd. means for determining the temperature of said block in a first sample interval, wherein said first sample interval is an interval of time designated as time n;
wherein said computing apparatus includes means for determining the temperature of said liquid sample mixture as a function of the temperature of said sample block over time by utilizing the relationship;
space="preserve" listing-type="equation">Tsamp.sub.n =Tsamp.sub.n-1 +TB.sub.n -Tsamp.sub.n-1) * tinterval/tauwhere Tsampn is equal to the sample temperature in said first sample interval, Tsampn-1 is a sample temperature in a second sample interval immediately preceding the first sample interval, said second sample interval designated as time n-1 , TBn is equal to the block termperature in said first sample interval, tinterval is a time in seconds between consecutive sample intervals, and tau is a function of thermal characteristics of said apparatus.
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Accused Products
Abstract
An instrument for performing highly accurate PCR employing a sample block in microtiter tray format. The sample block has local balance and local symmetry. A three zone film heater controlled by a computer and ramp cooling solenoid valves also controlled by the computer for gating coolant flow through the block controls the block temperature. Constant bias cooling is used for small changes. Sample temperature is calculated instead of measured. A platen deforms plastic caps to apply a minimum acceptable threshold force for seating the tubes and thermally isolates them. A cover isolates the block. The control software includes diagnostics. An install program tests and characterizes the instrument. A new user interface is used. Disposable, multipiece plastic microtiter trays to give individual freedom to sample tubes are taught.
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Citations
167 Claims
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1. An apparatus for controlled automated performance of polymerase chain reactions in at least one sample tube containing a known volume of a liquid sample mixture, which apparatus comprises:
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a. A sample block having at least one well for said at least one sample tube, b. a computing apparatus, c. heating and cooling means controlled by said computing apparatus for changing the temperature of said sample block, and d. means for determining the temperature of said block in a first sample interval, wherein said first sample interval is an interval of time designated as time n; wherein said computing apparatus includes means for determining the temperature of said liquid sample mixture as a function of the temperature of said sample block over time by utilizing the relationship;
space="preserve" listing-type="equation">Tsamp.sub.n =Tsamp.sub.n-1 +TB.sub.n -Tsamp.sub.n-1) * tinterval/tauwhere Tsampn is equal to the sample temperature in said first sample interval, Tsampn-1 is a sample temperature in a second sample interval immediately preceding the first sample interval, said second sample interval designated as time n-1 , TBn is equal to the block termperature in said first sample interval, tinterval is a time in seconds between consecutive sample intervals, and tau is a function of thermal characteristics of said apparatus. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 158, 159)
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124. Thermocycler apparatus for automated performance of a polymerase chain reaction comprising
a. a metal sample block having a major top surface and a major bottom surface, b. an array of spaced-apart sample wells formed in said major top surface, c. bias cooling constantly applied to said sample block at a rate sufficient to cause said block, if at a temperature within the range of 35° - -100°
C., to cool uniformly at a rate of at least about 0.1°
C./sec unless external heat is supplied, andd. computer-controllable heating means for uniformly raising the temperature of said block at a rate greater than the bias cooling rate, said thermocycler apparatus comprising means for maintaining the array of sample wells at a constant in the range of 35°
-100°
C. within a tolerance band of plus or minus about 0.5°
C. - View Dependent Claims (125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147)
- -100°
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148. Thermocycler apparatus for automated, rapid performance of a polymerase chain reaction comprising
a. a thermally homogeneous metal sample block of low thermal mass having a major top surface and a major bottom surface, said block containing in a central region of its upper surface an 8-by-12 rectangular array of sample wells having center-to-center spacing compatible with industry standard microtiter plate format, said block also containing a peripheral region surrounding said array, said peripheral region comprising a guard band having thermal characteristics similar to the thermal characteristics of the central region, b. a bias cooling system for constantly cooling said sample block at a bias cooling rate sufficient to cause said block, if at a temperature within the range of 35° - -100°
C., to cool uniformly at a rate of at least about 0.1°
C./sec unless external heat is supplied,c. a computer system for receiving and storing user data regarding times and temperatures defining a plurality of reaction cycles, d. a ramp cooling system controlled by said computer system for selectively cooling said sample block at a ramp cooling rate of at least about 4°
C./sec from 100°
C. and at least about 2°
C./sec from 40°
C.,e. a multizone heating system controlled by said computer system having a heating zone for the central region of the block and a heating zone for the guard band, said heating system comprising means for providing heat necessary to maintain the sample block at a constant temperature in the range of 35°
-100°
C. and also capable of providing ramp heating to the block,f. a pressing cover vertically displaceable above said sample block, and g. cover displacing means for raising said cover and for lowering said cover and maintaining its vertical position against a resisting force of at least about 3000 grams, said thermocycler apparatus comprising means for maintaining the array of sample wells at a constant temperature in the range of 35°
-100°
C. within a tolerance band of plus or minus 0.5°
C. - View Dependent Claims (149, 150, 151, 152, 153, 154, 155, 156, 157)
- -100°
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160. In a combination of a thermal cycler for performing a polymerase chain reaction comprising a temperature-controlled metal block having an array of tapered wells in its top surface, and a plurality of individual reaction tubes having similarly tapered lower sections and upper sections which project above the top surface of the block when the tubes are placed in said wells, the improvement comprising means for seating said tubes in said wells, said seating means comprising:
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a) resiliently deformable sealing caps removably attached to said tubes, b) a platen, c) support means for supporting the platen above the block, and d) displacement means associated with the support means for raising the platen above said caps and for lowering it so as to apply a force of at least 30 grams to each cap. - View Dependent Claims (161, 162, 163, 164, 165, 166, 167)
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Specification