The present invention relates to the field of analyzing biological probes and particularly relates to an apparatus and method for analyzing multiple subsets of probes emitting light, for example fluorescent and bioluminescent probes.
Biological probes have been designed to identify a particular biological substance by attaching to that substance. In use, the probes are attached to a substrate, and an unknown material is treated with markers or tags before applying the unknown material to the probes. Often, a fluorescent tag is attached to the unknown material, and the material is exposed to the probes. If the unknown material includes a particular type of bacteria, and if a probe for those bacteria is present, the bacteria will attach to the probe. Then, the tag carried by the bacteria may be detected. Examples of detectors for biological probes are described in the U.S. Pat. Nos. 6,197,503 and 6,448,064. Another technique attaches biological probes that include fluorescent material that will emit light only if the probe is exposed to its particular target material.
Expense is a primary stumbling block for applying this technology to a broad base of industrial and commercial applications. Often, highly skilled technicians are required to prepare the slide, appropriately expose it to a material in question, and read the slide. Typically, the slides are expensive and the machines used to read the slides are even more expensive. Thus a need exists for an inexpensive fast mechanism and method for reading biological slides by trained personnel who are not necessarily highly skilled in the biological field.
The present invention provides an apparatus and method for economically producing, using and reading biological slides, meaning slides or other support structure carrying biological probes. The technology of the present invention will lower the cost of using biological slides to the point that everyday commercial applications will be possible. For example, restaurants and groceries may routinely check their meat and other foods for specific types of bacteria.
In accordance with present invention, an analyzer is provided for reading biological probes. The analyzer includes a housing, and a slide carriage is mounted for movement within the housing. A slide is positioned on the slide carriage, and a plurality of probes are disposed on the slide in a defined pattern. Each probe may generate an electromagnetic probe signal, to indicate prior exposure to a predetermined substance. Alternatively, the probe signal may not be produced, which indicates the absence of prior exposure to a predetermined substance. The detector is configured to detect probes signals from a plurality of probes located in a defined pattern on the slide. Also, the defined pattern is denser than the first pattern of the detector and is configured in a shape corresponding to a plurality of first patterns so that the detector can sense multiple subsets of the probes within a probe pattern on the slide.
In a preferred embodiment the detector includes a plurality of sensors arranged in the first pattern, and each sensor is disposed for detecting a probe signal from a single probe set. The plurality of sensors on the detector is aligned with a subset of the probes on the slide when the slide is being read. Preferably the sensors are arranged on the detector in a first pattern and the probe pattern on the slide is arranged in the form of interlaced first patterns. Thus, the detector may be aligned with a first pattern of sensors aligned with a first pattern of probes constituting a subset of the probe pattern. All of the probes on the slide may be read by sequentially moving the detector or the slide or both, one relative to the other, from one subset to another subset of probes on the slide. In one embodiment the first pattern and the probe pattern both comprise a plurality of locations arranged in rows and columns. The number of rows and columns in the probe pattern is a multiple of the number of rows and columns in the first pattern. For example, if the first pattern has 5 rows, the probe pattern may have 10 rows or 15 rows, etc.
In another embodiment the first pattern is in the shape of a pie section and the probe pattern is arranged on the slide in a radial configuration about a center point. Again, the probe pattern preferably is denser than the first pattern, and most preferably the probe pattern includes multiples of the first pattern. With the probe pattern arranged about a center point on the slide, the slide may be read by aligning the detector with one first pattern on the slide, reading the probes with which the detector is aligned, relatively moving the slide and the detector to align the detector with subsequent sets of probes on the slide arranged in the first pattern, reading the second set of probes, and continuing to relatively move the detector and slide and read additional probes, preferably until all probes on the slide are read.
In accordance with another aspect of the invention, the analyzer includes a source of electromagnetic radiation (preferably a scanning laser) for illuminating the probes at selected times and the source is extinguished at other times so that the probes are either illuminated or not. The probes are preferably constructed in part from a fluorescent material to indicate exposure to a selected material. Thus, certain probes emit a probe signal in the form of fluorescent light after being illuminated by the source and other probes do not. In one embodiment, the probes that include a fluorescent material were made by first placing a material at the probe location that will bind to another specific material, such as nucleotides or DNA. Then, an unknown material has fluorescent markers attached to it, and then both the unknown material and the fluorescent markers or tags are exposed to the probes. If the unknown material is the material to which the probe binds, then the unknown material will bind to the probe and the fluorescent marker will be present on the probe on the slide. If the unknown material does not bind to a particular probe, then that particular probe will not include fluorescent material. Therefore, the presence or absence of fluorescent material on each probe indicates the presence or absence of particular types of material, such as a particular strain of bacteria, in the unknown material.
In an alternate embodiment, all of the probes have a fluorescent material attached to the probe, but the fluorescent material will not fluoresce unless and until the probe is exposed to the particular substance that a particular probe is designed to detect. Thus, if such particular probe fluoresces when exposed to the source, it means the probe was exposed to a particular substance. If such probe does not fluoresce when exposed to the electromagnetic radiation of the source, it means that the probe has not been exposed to the particular substance, and the fluorescent material will not fluoresce even though it resides on the probe and has been illuminated.
In order to align the detector with the slide, alignment indicia is preferably disposed on the slides to enable proper alignment. Preferably one or more indicia are placed on the slide in the location of one or more probes. The indicium produces a probe signal in the form of light that is detected by the sensors. The detector aligns itself by aligning the detector sensors over the indicia in the probe locations. Preferably the drive mechanism for the slide carriage includes an X, Y, Z, and R drive mechanism for moving of the slide carriage in X, Y and Z directions, which are non-parallel directions, preferably orthogonal directions. Also, the drive mechanism rotates the slide carriage about an axis, “R”. Using the drive mechanism, the detector is moved until the expected light from the alignment indicia is received by the expected sensors. Since the alignment indicia is controlled to contain a specific amount of light producing material that produces a specific amount of light, and since the exact position of the alignment indicia is known, it is also known that the alignment indicia will produce a specific reading (the detection signal) in specific sensors when the detector is properly aligned over the slide.
Thus, by moving the detector with respect to the slide until the expected specific amount of light is received in the known specific sensors, the slide is aligned with the detector. (As used herein, mechanism and mechanisms are synonymous, and either may have many parts or just one part.)
Additional aspects and advantages of the present invention may be understood by reference to the following Detailed Description when considered in conjunction with the attached drawings in which:
Referring now to the drawings in which like reference characters designate like or corresponding parts throughout the several views, there is shown in
The slide 28 has been prepared in advance before it is placed into the analyzer 20. For example, a slide may be prepared with a plurality of the probes formed on the slide with each probe designed to bind with a specific type of biological material. In one embodiment, the biological probes are constructed using a detector material with a fluorescing material associated with the detector material. Each probe may have different detector material that reacts or detects different substances. The fluorescing material is associated with the detector material such that the fluorescing material transmits fluorescent light from the probe only when the detector material has contacted the particular substance it is designed to detect. Thus, if the particular substance is present, the probe will emit fluorescent light when illuminated by an appropriate source of electromagnetic illumination. For example, the probe may be designed to fluoresce when a particular food bacteria is exposed to the probe.
In an alternate embodiment, the probes are designed to detect and attach to particular substances, but the fluorescing material is attached to the target bacteria, not the probe. A material of interest, such as a food product, is then treated with fluorescent markers that attach to the food product and extraneous substances that may be present in the food products. For example, if bacteria are present in the food product, the fluorescent markers attach to the bacteria. The treated material of interest is then exposed to the slide 28. If one of the probes is designed to bind to salmonella bacteria, and if salmonella is present in the food product, the salmonella bacteria will bind to that particular probe of the slide 28. The salmonella bacteria will carry the fluorescent markers and, thus, the presence of the salmonella on the probe may be detected by detecting fluorescent light from the fluorescent markers on the salmonella, which is bound to the probe. This one example illustrates the many types of slides of that may be prepared for the analyzer 20. A common feature of each slide is that it includes a plurality of spots that will produce signals, such as light, and thereby provide information.
Referring to
A more detailed view of the slide carriage 32 is shown schematically in
The rotational drive mechanism 37 is configured to rotate the carriage 32 through 360 degrees of rotation in either direction as indicated by drive train 41. The rotational drive mechanism 37 includes an appropriate motive mechanism such as drive motor 38. Operating above or on top of the rotational drive mechanism 37 is a vertical drive mechanism 40 that drives the carriage up and down in the vertical direction, the Z direction. Again, the vertical drive mechanism 40 includes a motor 42 and a drive train 43, such as a drive screw, chain or belt. Drive motor 42 provides force to move the mechanism 40 and thereby move the carriage 32 up and down. On top of the vertical drive mechanism 40, a horizontal Y drive mechanism 44 is mounted and includes a drive motor 49 and a drive train 47. The horizontal Y drive mechanism 44 moves the carriage 32 horizontally in the Y direction. Finally, a horizontal X drive mechanism 48 is mounted above the horizontal Y drive mechanism 44. The drive mechanism 48 moves the carriage 32 in the X direction as indicated in
While it is preferred to provide the movement of the slide 28 in the manner previously described, in alternate embodiments, different types of movement may be provided. For example, in some applications, it may be desirable to provide three-dimensional movements along non-perpendicular directions. Also, a different order of movement might be desirable in other embodiments. In the above example, rotational movement caused by drive mechanism 37 is the base form of movement upon which all other movements are built. In alternate embodiments, the rotational movement could be provided as the highest order of movement. That is, the rotational drive mechanism 37 would be mounted on top of the other portion of the drive mechanism. In the embodiment shown in
Also, the movement of the slide 28 is relative to the detector 56 (
Referring now to
The conditioning optics 57 may include lenses and filters. Preferably the conditioning optics 57 will filter ambient light and light at the frequency of the light applied to the slide 28, such as the frequency of light rays 63, 62 and 65. Thus, conditioning optics 57 is designed to filter out the light rays 63, 62 and 67 preventing it from reaching the detector 56.
A view of the top side of detector 56 is schematically shown
The detector 56 in this embodiment includes an array of twenty-five sensors 58 arranged in five rows and five columns. The row and column numbers are labeled along the left and top side of the detector 56, respectively. For reference purposes the top left sensor 58 is positioned at a row 1, column 1 and therefore has a position of 1, 1. For convenient reference purposes, this sensor will be labeled sensor 58-1,1. The sensors 58 are formed on a substrate 59 that is preferably constructed of a semiconductor material, such as silicon. The sensors 58 are each light sensitive photo diodes that a produce an electrical signal in response to receiving light. In this particular embodiment, the sensors 58 are preferably round having a diameter 60 of about 1.0 mm. The sensors in this embodiment are separated in the Y direction by a Y distance 64 equaling about 0.5 mm and are separated in the X direction by an X distance 66 equaling 0.5 mm. These precise dimensions are given for purposes of illustration only are not intended to limit the scope of the invention. Also, the detectors 58 may be rectangular, oval, or other shapes.
Referring to
To read the array 70, the slide 28 is moved until the detector 56 is positioned over the array 70 with sensor 58-1, 1 positioned over the probe labeled “A”. The array of sensors 58 is aligned with the probes 78 such that each sensor 58 will read an “A” probe 78 when the sensor 58-1, 1 is positioned directly over the sensor 78 labeled “A” in the upper left-hand corner of array 70. After all of the “A” probes 78 are read, the slide 28 moves to the left and, in a relative sense, the detector 56 moves to the right and positions sensor 58-1, 1 over the “B” probes 78 in the upper left-hand corner of array 70. Likewise, all other sensors 58 are positioned over “B” probes 78 in the array 70. After the “B” probes are read, the slide 28 moves upwardly, and in a relative sense, the detector 56 moves downwardly in the Y direction and positions the sensor 58-1, 1 over the “C” probe 78 in the upper left-hand corner of the array 70. Again, all of the other sensors 58 are positioned over the other “C” probes 78 in the array 70 and the “C” probes are read. Finally, the slide 28 moves right and, in a relative sense, the array of sensors 58 are moved to the left positioning sensor 58-1, 1 over the “D” probes in the upper left-hand corner of the array 70. The other sensors 58 are positioned over the other “D” probes 78 and the “D” probes are read. In this embodiment, only the slide 28 moves, but in other embodiments, the detector 56 may be constructed to move, or both the detector 56 and slide 28 may move.
After array 70 and has been completely read, the detector 56 is positioned over the array 72 and the reading process is repeated. Thereafter, arrays 74 and 76 are read in the same manner. In
Referring again to
The positions of the probes 78 on the slide 28 are very precise and the positions of the sensors 58 on the detector 56 are very precise, but the relative positioning between the slide 28 and the detector 56 is less precise because the slide 28 must be removed and moved during operation of the analyzer 20. In practice, once the analyzer 20 is calibrated properly, the detector 56 and the slide 28 may be accurately and properly positioned without alignment checks. However, in the preferred embodiment, alignment checks are provided. Preferably on every slide, the first and the last “A” probes 78 are seeded with material that will fluoresce brightly. For example, referring to array 70 in
Referring now to
The irregular pattern of sensors 82 and the detector 80 are configured to read the irregular pattern of probes 96 disposed on the slide 86 shown in
Referring to
Referring again to
The patterns in each of the arrays 112-116 are oriented in a radial direction with respect to the center 118 of the circular slide 102. A radial configuration does not require that any row of probes 110 be radial with respect to the center 118; it only requires that the patterns be oriented on the slide 102 such that the patterns of probes may be aligned with the pattern on the detector(s) by rotation about the center 118, plus linear X-Y movement. That is, all of the patterns substantially point to the center 118 of the slide 102. Thus, the slide 102 may be read by a combination of rotating the slide 102 and shifting the slide 102 in the X and/or Y direction. To read the slide 102, the detector 100 is first positioned over the array 112 with the sensors 104 positioned over the probes 110-1 (illustrated by the white dots). After the probes 110-1 are read, the slide 102 is rotated counterclockwise until the detectors 104 align with the probes 110-2 (illustrated by black dots). After the probes 110-2 are read, the slide 102 must be rotated slightly in a clockwise direction and the slide must be shifted to the left until the probes 110-3 are aligned with the detectors 110. When shifting between subsets of probes 110, the slide may also be moved in a linear direction, such as, in the X and Y direction with no rotation. That is, the slide 102 could be moved to the left and up to align the sensors 104 with the probes 110. Of course, the patterns 110 must be positioned to allow such movement, and in such case at least one pattern will not be oriented radially with respect to center point 118. This linear movement may be accomplished using the X and Y drive mechanisms 48 and 44, respectively.
After the array 112 has been read, the slide 12 is rotated in a counterclockwise direction and it is shifted in the X and Y directions until the probes 110-1 (the white dots) of array 114 are aligned with the sensors 104 of the detector 100. Then, the slide 102 is rotated or shifted in the X, Y direction, or both, to read all three patterns in the array 114 in the same manner as previously described. In like manner, arrays 115 and 116 are read.
The arrays and patterns illustrated in
In the illustration of
Referring now to
After the probe 132 has been illuminated, the external light sources are extinguished and fluorescent light is produced by the probe 134 as illustrated by light rays 144. The lens 136 may be greater in diameter than the well 132 and extend partially around the probe 134 so that the lens 136 collects light emanating from the probe in multiple directions and directs the light to the detector 138 as indicated by the light rays 146.
Preferably, the detector 138 is positioned in the image plane of the lens 136 to maximize the energy that is collected and concentrated on a detector 138. In addition, conditioning optics 148 may be provided between the detector 138 and the probe 134. The conditioning optics 148 preferably includes a filter that blocks light having a frequency ambient light or the illumination light as represented by light rays 140, 142. The filter of the conditioning optics 148 will transmit light having a frequency equal to that of the fluorescent light emanating from the probe 134 as illustrated by light rays 144. Optics 148 preferably also include a lens or lenses that directs light to the detector 138, such as by producing an image of the probe 134 on the detector 138.
Referring now to
After exposing the probe 153 to illuminating light such as light rays 152 and 154, fluorescent light is produced by the probe 153 as represented by light rays 156, 158 and 160. Some of the fluorescent light will project directly toward the detector 138 and strike the face of the detector 138, penetrating the detector 138. Fluorescent light as illustrated by light rays 158 and 160 will be transmitted by the probe 153 in the direction of the walls of the well 151. Upon striking the well 151, the light will be redirected toward the detector 138 as indicated by the light rays 162 and 164 representing light rays reflected from the well 151. The well 151 has a parabolic shape, or a modified parabolic shape designed to maximize the amount of fluorescent light that is reflected toward the detector 138. In practice, the parabolic shape of the well 151 is dictated by the position of the top surface of the probe 153. The parabolic shape of the well 151 is configured to maximize the amount of light that is reflected onto the detector 138, where the light is transmitted from the top surface of the probe 153 in substantially all upward directions.
Conditioning optics 166 are also provided to further condition of the fluorescent light emitted by the probe 153 and the illumination light represented by light rays 152 and 154. For example, in a preferred embodiment, the conditioning optics 166 would include a filter designed to totally block light having a frequency of the illumination light rays 154 and 152, but the filter would totally pass light having a frequency of the fluorescent light emitted by the probe 153, such as light rays 156, 158 and 164. By collecting and focusing the fluorescent light emitted by the probe of 143, the well 151 is functioning in a manner analogous to the lens 136 shown in
Referring now to
The internal operations of the analyzer 20 as described above may be understood at a more detailed level by reference to
The data processor 196 is also connected to monitor and control the cassette drive mechanism 36. When the user desires to insert a new slide into the analyzer 20, the command to open the cassette 24 is entered by the user through the keypad 26. The data processor 196 then issues commands to the cassette drive mechanism 36 causing it to extend the cassette 24 out of the housing 22. After a slide 28 has been inserted onto the cassette 24, the user inputs commands through the keypad 26, and in response to the user commands, the data processor 196 issues commands and controls the cassette drive mechanism 36 to translate the cassette 24 into the housing 22 and adjacent to the detector 56. In one embodiment, the slide 28 is read immediately when the slide 28 is placed on the cassette 24 and the cassette is withdrawn into the housing 22. In other embodiments, the data processor 196 will await commands from the user through keypad 26 before it begins of the process of reading the slides 28.
To begin the reading process, the data processor 196 issues commands to the X Y Z and R drive mechanisms 37, 40, 44, and 48. In response to these commands, the detector, such as detector 56 shown in
After the slide and detector have been aligned, the data processor 196 issues commands to turn on a source of light, such as a laser 204 shown in
After the entire slide has been read, the slide 28 may be removed from the analyzer 20 in response to commands issued by the user through the keypad 26, or the data processor 196 may be programmed to automatically eject the slide after it is read. The data that was obtained by the data processor 196 is initially stored in memory 198. However, under the control of the user 26, the data may be exported from the data processor 196 through such devices as a printer 202, “the display 30” or an electronic port 206 that communicates with other data processors and data collection devices. While specific examples of the invention have been discussed above, it will be understood that these examples are intended for the purpose of illustration only. It will be understood that the present intention is capable of numerous arrangements, modifications and substitutions of parts without departing from the scope and spirit of the invention as defined by the appended claims.