Chemical apparatus and process disinfecting – deodorizing – preser – Control element responsive to a sensed operating condition
Reexamination Certificate
1996-01-16
2003-12-09
Alexander, Lyle A. (Department: 1743)
Chemical apparatus and process disinfecting, deodorizing, preser
Control element responsive to a sensed operating condition
C422S063000
Reexamination Certificate
active
06660233
ABSTRACT:
TECHNICAL FIELD
This invention relates to a system and methods for detecting the presence of target biomolecules within samples with robotic assistance for a sample holder carrying an array of reactants.
BACKGROUND ART
Assays for the detection of target biomolecules within a sample, especially of multiple target biomolecules within a sample, are often performed by applying a volume of the sample to a test slide, membrane, or other substrate having immobilized reactants which may interact with the target or targets to form detectable complexes. These immobilized reactants are usually disposed at fixed locations, with samples brought to these locations. U.S. Pat. No. 5,139,743, for example, discloses a. biochemical analysis apparatus wherein an applicator takes up a liquid sample and applies the sample to a fixed position test film for chemical analysis of the sample.
Sometimes complexes of target biomolecules and reactants are visually detectable directly after an appropriate incubation period for the sample and reactants, or after numerous development steps wherein development chemicals, such as fluorescent dye-conjugated molecules, are allowed to interact with the complexes. For example, the detection mechanism in U.S. Pat. No. 5,296,194 involves optically detecting a color change in a blood drop applied to a test slide.
U.S. Pat. No. 4,877,745 discloses methods for preparing immobilized reagents and applying samples to immobilized reagents. In particular, this patent discloses dispensing precisely controlled volumes of droplets onto a medium at precisely controlled locations, to form arrays of immobilized reagents by a jet head. An x-y plotter may be modified to carry a jet head so that reagent may be dispensed over an area.
Robotic laboratory workstations, such as the Biomek 1000 and 2000 of Beckman Instruments, Inc. have been developed for automatically carrying out assays involving multiple reactants and multiple samples. Typically such workstations are designed to deliver robotically precise volumes of reactants to a number of different samples located at known areas within the workstation. Alternatively, workstations can robotically move samples to reagents.
U.S. Pat. No. 5,171,537 to Wainwright et al. teaches activated immunodiagnostic pipette tips. The pipette tip houses a spherical element which is coated with a single ligand having affinity for a target molecule of a sample. With this device, the test element may be brought to contact the sample, as by aspirating the sample into the pipette tip. These pipette tips are limited in their sample throughput because they house only a single ligand reagent and thus preclude the detection of multiple analytes within a sample.
A class of devices known as optical biosensors, characterized by immobilized assay species within a supporter and a light collection device coupled to an optical waveguide, is also known. Optical biosensors may be used for detecting and quantifying the presence of specific species in test fluid samples, such as in clinical diagnostic reactions. For example, U.S. Pat. No. 4,857,273 discloses an optical biosensor for immunoassays and certain other reactions. Other examples, involving use of an optical fiber, are U.S. Pat. No. 5,143,066 and U.S. Pat. No. 5,401,469.
It is an object of the present invention to provide apparatus and methods for rapidly and automatically determining the presence of multiple target biomolecules in a single sample. It is another object of the present invention to provide analytical methods which require minimal sample volume and a minimal number of liquid transfers. It is a further object of the present invention to provide a device and system for rapid assessment of samples for target biomolecules which is readily adaptable to a variety of chemical and other detection schemes.
DISCLOSURE OF THE INVENTION
The present invention achieves the above objects by providing an analytical biochemistry system for automated analysis of samples for the presence of target biomolecules. The system includes a solid substrate which is supported by a holder and carried by a manipulator, such as a robotic arm. Immobilized on the solid substrate surface at discrete, site-specific locations are reactants in an array which are capable of binding with target biomolecules in specific binding reactions to form immobilized biomolecule complexes. Such an array is termed a “bioarray”. The presence of target biomolecules in the sample is determined by detecting immobilized biomolecule complexes on the bioarray with some kind of probe, e.g. a fluorescence detector. In operation, the manipulator moves the bioarray to contact the substrate surface with a volume of sample. Then the manipulator moves the contacted bioarray to a detection station to detect the absence or presence of immobilized biomolecule complexes. In alternative embodiments the bioarray is stationary and a sample manipulator moves samples to the holder.
In the preferred embodiment, the bioarray is mobile, being carried by a manipulator. A detection station is located near the sample to probe the substrate after interaction between the reactants and sample or samples has occurred.
Distinct reactants specific to different target biomolecules are immobilized on a preferably flat, nonporous substrate. These reactants form a plurality of active sites on the substrate at known locations. The substrate may be a planar strip with linearly-arranged reactants forming separable spots or bands, or may be a planar sheet having an area-wide arrangement of reactants, forming spots or dots in a two-dimensional array, or may be a fiber or rod with substrate disposed in a manner similar to a strip.
The holder supports the bioarray and is carried by the manipulator which transports the substrate to the location of the fixed sample, and then to the location of the detection assembly. As stated, the substrate could be fixed and the sample transported. One example of a holder is a pipette or a pipette tip, within which a bioarray is affixed. The sample is drawn up into the pipette tip, as with aspiration from a bulb or vacuum pump, or withdrawal of a plunger. The sample is thus placed in contact with the substrate, allowing any target molecules which may be present within the sample to interact with the appropriate reactive sites on the substrate. After the appropriate incubation or reaction period, the sample may be removed from the pipette tip, as by air pressure or positive displacement with a plunger.
Another example of a useful holder is a pipette adapter resembling a truncated pipette tip and having a bracket or a flat surface for supporting the substrate. The pipette adapter may be placed directly into a sample, such as in a well of a microtiter plate or in a vial, in order to provide contact of the holder and the sample. The pipette adapter and accompanying substrate are then removed from the sample to a detector station. The various holders of the present invention may be adaptations of standard pipetting tools. The holders also are designed to require minimal sample volumes and to allow optical inspection of the substrate with minimal interference by the holder.
The method for detecting target biomolecules within a sample includes the steps of treating a substrate with a plurality of distinct reactants to form reagents immobilized on the substrate at fixed, known positions defining an array, i.e. a bioarray. The reactants are selected to bind one or more target molecules to form a complex having a detectable and identifiable characteristic, such as a fluorescent signature. The bioarray is supported in the holder. In turn, the holder has a shape which can be picked up by a manipulator which moves the substrate for contact with the fixed sample, and then removes and possibly rinses the substrate at another location to remove unbound biomolecules. Then the manipulator moves the substrate to a probing station, such as an optical inspection location for probing the active sites of the substrate with a beam for determining complementation of the target biomo
Coassin Peter J.
Helphrey David B.
McNeal Jack D.
Alexander Lyle A.
Beckman Coulter Inc.
Hill D. David
May William H.
Sheldon & Mak
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