United States Patent [191
`Fernandes et a1.
`
`[54] MUTLI-FUNCTIONAL PHOTOMETER
`WITH MOVABLE LINKAGE FOR RQUTING
`OPTICAL FIBERS
`
`[75] Inventors: Jorge Fernandes, San Francisco;
`Michael C, Norris, Los Gatos’ both
`of Calif
`
`[73] Assignee: Biolumin Corporation, San Jose,
`Calif.
`
`[21] Appl. No.: 100,541
`
`[22] Filed:
`
`Jul. 30, 1993
`
`[51] Int. Cl.9 ................... .. G01N 21/59; G01N 21/64;
`[57] U 5 C1
`356/7gO32512/63/1284:
`“
`'
`'
`‘ ""
`56/436; 356/440;
`250/4581; 385/25
`[58] Field of Search ............... .. 356/73, 317, 318, 319,
`356/326, 328, 402, 409, 410, 414-419, 432,
`434-436, 440; 250/227.23, 227.26, 458.1, 459.1,
`461.1, 461.2; 385/15, 16, 18, 19, 25
`_
`References Cited
`U.S, PATENT DOCUMENTS
`3,697,185 10/1972 Kassel et a1, ...................... .. 356/410
`3_874_78O 4/1975 Love _
`4373779 2/1983 Dorsey _
`4,477,190 10/1984 Liston et a1_ ______________________ ,_ 356/413
`4,501,970 2/1985 Nelson ............................ .. 250/4581
`4,587,812 5/1986 Brega .
`4-6Zl463 11/1986 Stefanski e1 31- --------------- -- 250/4581
`
`[56]
`
`4.626,684 12/1986 Landa . . . . 1 . . , . , , .
`
`. . . . . .. 356/318
`
`396/31?
`41699338 9/1987 Me‘cr
`' ' ' "1356/ 73
`4’730’9I2 3/1988 Béich et a1‘ ' ' ‘ ' '
`556/417
`4,750,857 6/1988 Glfford ct al.
`250/4581
`4’799‘756 1/1989 Hirschfeld
`4,802,768 2/1989 Gifford et al. .................... .. 356/417
`4,815,812 3/1929 Miller .
`4,820,045 4/1989 Boisde et a1. ..................... .. 356/319
`
`46
`
`lllllllllllllllllllllllllllllllllll
`I|||||Illllllllllllllllllllllllllllll
`USOO5436718A
`Patent Number:
`5,436,718
`[11]
`[45] Date of Patent:
`Jul. 25, 1995
`
`4,840,485 6/1989 Gratton ............................ .. 356/317
`4,937,457 6/1990 Mitchell .
`250/4581
`4,945,245 7/1990 Levin ....... ..
`250/4581
`4968,1423 11/1990 Chow et a1.
`356/427
`5,030,832 7/1991 Williams et a1. ..
`250/4581
`5,125,747 6/1992 Sayegh et all ...... 1.
`356/407
`5,131,746 7/1992 O’Rourke et a1.
`356/319
`5,141,609 8/1992 Sweedler et al.
`356/344
`5,143,853 9/1992 Walt ................ ..
`436/501
`5.151.869 9/1992 Alcala ............................ .. 250/4581
`
`FOREIGN PATENT DOCUMENTS
`
`62160 10/1982 European Pat. Off. .......... .. 356/414
`
`OTHER PUBLICATIONS
`“The Cytoflour Flourescence Measurement System".
`Automated Flourescence Scanning, Millipore, 1990.
`Primary Examiner—F. L, Evans
`Attorney, Agent, 01' Firm—Blake1y, SOkOIOff, Taylor &
`Zafman
`
`ABSTRACT
`[57]
`A rnulti-functional photometer includes a scanning
`mechanism having a housing (10) that bears a movable
`linkage (12). The linkage is coupled to an optical scan
`“i119. head (.18) and incorporates Optical ?bers .for trans‘
`mittmg radiant energy to and from the scanmng head.
`The arm comprises a C-shaped “elbow” member (14),
`pivotally attached to a “shoulder” member (16). In turn,
`the “shoulder" member of the arm is pivotally con
`nected to the housing. Dynamic couplings join the opti
`cal ?bers such that the shapes thereof remain ?xed re
`
`gardless of the orientation of the arm. The housing
`
`further incorporates a Cartesian-coordinate table (20)
`for positioning the scanning head with respect to 21
`.
`.
`.
`_
`microplate (22) that contams a plurality of analyte sam
`P199
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`47 Claims, 10 Drawing Sheets
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`THERMO FISHER EX. 1031
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`U.S. Patent
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`July 25, 1995
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`Sheet 1 of 10
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`US. Patent
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`July 25, 1995
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`Sheet 2 of 10
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`All “
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`U.S. Patent
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`July 25, 1995
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`Sheet 3 of 10
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`July 25, 1995
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`Sheet 4 of 10
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`US. Patent
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`July 25, 1995
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`Sheet 5 of 10
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`FIG.5
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`US. Patent
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`July 25, 1995
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`Sheet 6 of 10
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`US. Patent
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`July 25, 1995
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`Sheet 7 0f 10
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`I32
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`I54
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`? 54
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`FIG.6
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`I54
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`FIG.7
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`U.S. Patent
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`July 25, 1995
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`Sheet 8 0f 10
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`FIG.8
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`US. Patent
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`July 25, 1995
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`Sheet 9 of 10
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`US. Patent
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`July 25, 1995
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`Sheet 10 0f 10
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`5,436,718
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`START
`
`ZOO
`N/
`IDLE LIGHT
`SOURCE
`
`II
`
`NO
`
`'
`
`202
`,/
`
`OPERATING
`INSTRUCTIONS
`RECEIVED ?
`
`204
`
`Ives
`SELECT
`SPECIFIED
`BANDPASS
`FILTERS
`
`POWER'UP
`LIGHT
`SOURCE
`
`/208
`
`MOVE
`SCANNING
`HEAD I'HOME'I
`AND CALIBRATE
`PHOTODETECTORS
`
`YES
`
`1
`
`SCAN
`SAMPLES
`
`REPEAT
`SCAN ?
`
`FIG.IO
`
`THERMO FISHER EX. 1031
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`

`
`1
`
`MUTLI-FUNCI‘IONAL PHOTOMETER WITH
`MOVABLE LINKAGE FOR ROUTING OPTICAL
`FIBERS
`
`FIELD OF THE INVENTION
`The present invention relates to the ?eld of spectros
`copy, particularly to a multi-functional photometer
`capable of measuring light absorbance, ?uorescence,
`and luminescence of a sample.
`
`10
`
`5,436,718
`2
`croplates are bene?cial since they allow simultaneous
`preparation of a large number of test samples. More
`over, microplates are inexpensive, safe, sturdy, and
`convenient to handle. They are also disposable and can
`be cleaned easily when necessary.
`One instrument currently available for ?uorescent
`analysis of samples in microplate wells is the Cyto?uor
`2300 ?uorometer, distributed by Millipore Corporation,
`Bedford, Mass. This ?uorometer includes a scanning
`head that resides underneath the microplate and moves
`along the bottom face thereof to scan the sample sites.
`The scanning head interfaces with the optical system of
`the device via a bundle of optical ?bers that transmits
`excitation and emission radiation.
`However, the capabilities of the Cyto?uor 2300 ?u
`orometer are limited in that it cannot perform absor
`bance measurements. Furthermore, the movement of
`the scanning head from one microplate well to another
`continuously alters the geometrical con?guration of the
`optical-?ber bundle that is attached to the head. Conse
`quently, curvatures of the light-transmitting ?bers
`change, introducing variations in their optical proper
`ties. These variations create inconsistencies in readings
`between different wells and adversely affect the repeat
`ability, and thus, accuracy of measurements. Moreover,
`continuous bending of the ?bers produces stresses that
`cause mechanical failure of the ?ber cores.
`Additionally, to allow unrestricted movement of the
`scanning head, ?exible plastic ?bers are employed, as
`opposed to less pliable quartz ?bers. 0n the down side,
`plastic ?bers cannot efficiently transmit radiant energy
`in the ultraviolet (UV) region of the spectrum. Accord
`ingly, the ?uorometer is unable to perform measure
`ments, such as binding studies of certain proteins, e.g.,
`tryptophan, since ?uorescence analyses of this type
`require the use of UV radiation. Furthermore, the de
`formation resistance of the optical-?ber bundle slows
`the movements of the scanning head, thus limiting the
`ability of the apparatus to perform kinetic measure
`ments.
`Another spectroscopic apparatus utilizing micro
`plates is disclosed in US. Pat. No. 4,968,148 to Chow et
`al., 1990. Chow’s device uses an optical distributing
`element to selectively direct radiant energy to speci?ed
`microplate sites. One drawback of this instrument is its
`inability to perform fluorescence measurements. More
`over, the large number of ?bers unnecessarily compli
`cates the apparatus and increases production costs.
`Also, the light-delivery system of the instrument has a
`?xed geometry that can only accommodate a micro
`plate with one particular well layout. Chow’s apparatus
`does not have the versatility to be utilized with micro
`plates having different con?gurations of wells.
`
`BACKGROUND OF THE INVENTION
`In biological research, it is often necessary to assay
`samples for content of various chemicals, hormones,
`and enzymes. Spectroscopy, which is the measurement
`and interpretation of electromagnetic radiation ab
`sorbed or emitted when the molecules, or atoms, of a
`sample move from one energy state to another, is
`widely utilized for this purpose. Currently, the most
`common spectroscopic techniques pertain to measure
`ments of absorbance, ?uorescence, and luminescence.
`Chemical analyses with absorption spectroscopy
`allow one to determine concentrations of speci?c com
`ponents, to assay chemical reactions, and to identify
`individual compounds. Absorbance measurements are
`most commonly used to ?nd the concentration of a
`speci?c composition in a sample. According to Beer’s
`law, for a composition that absorbs light at a given
`wavelength, the total absorbed quantity of such light is
`related to the quantity of that composition in the sam
`ple.
`Fluorescence, in turn, is a physical phenomenon
`based upon the ability of some substances to absorb and
`subsequently emit electromagnetic radiation. The emit
`ted radiation has a lower_ energy level and a longer
`wavelength than the excitation radiation. Moreover, the
`absorption of light is wavelength dependent. In other
`words, a ?uorescent substance emits light only when
`the excitation radiation is in the particular excitation
`band (or bands) of that substance.
`For ?uorescence measurements, ?uorescent dyes
`called ?uorophores are often used to “tag” molecules of
`interest, or targets. After being irradiated by an excita
`tion beam, ?uorophores, bonded to the targets, emit
`light that is then collected and quantized. The ratio of
`45
`the intensity of the emitted ?uorescent light to the in
`tensity of the excitation light is called the “relative
`?uorescence intensity” and serves as an indicator of
`target concentration. Another useful characteristic is
`the phase relationship between the cyclic variations in
`the emitted light and the variations in the excitation
`light, i.e., the time lag between corresponding varia
`tions in the emission and excitation beams.
`As noted above, luminescence measurements can also
`be employed for analyzing biological samples. Lumines
`cence is the property of certain chemical substances to
`emit light as a result of a chemical change; no excitation
`from a light source is necessary. Moreover, lumines
`cence can~be produced by energy-transfer mechanisms
`that take energy of a high intensity, e.g., a radioactive
`emission, and transform it to energy of a low intensity,
`e.g., a ?ash of light.
`At the present time, a variety of spectroscopic instru
`ments is commonly used in the art. A number of these
`instruments are designed to be utilized in conjunction
`with multi-site analyte receptacles called “microplates”,
`which usually comprise one-piece structures having
`multiplicities of Wells for holding analyte samples. Mi
`
`65
`
`25
`
`55
`
`60
`
`OBJECTS AND SUMMARY OF THE
`INVENTION
`It is accordingly an object of the invention to provide
`a multifunctional photometer which overcomes the
`foregoing disadvantages, e.g., which measures absor
`bance, ?uorescence, and luminescence of a sample;
`which provides repeatable measurements and produces
`consistent readings between different test sites; which
`eliminates recurring bending of optical ?bers and me
`chanical failure thereof; which utilizes optical radiation
`ranging from the ultraviolet to the infrared spectrum;
`which is able to carry out kinetic measurements; which
`can accommodate microplates with different well con
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`?gurations; and which is relatively simple and inexpen
`sive to manufacture.
`Another object of the invention is to supply a pho
`tometer having a movable linkage for dynamically and
`interconnectingly routing optical ?bers such that a con
`stant con?guration thereof is always maintained during
`operation of the photometer.
`Yet another object of the invention is to provide a
`photometer which performs analyses of optical signals
`resulting from phenomena of absorbance, ?uorescence,
`and luminescence over a range of spectral wavelengths.
`Further objects and advantages will become apparent
`after consideration of the ensuing description and the
`accompanying drawings.
`In the preferred embodiment of the present invention,
`a multi-functional photometer includes a scanning
`mechanism having a housing that bears an articulated
`movable arm. The arm is coupled to an optical scanning
`head and incorporates light-transmitting conduits, such
`as optical ?bers, for transmitting radiant energy to and
`from the scanning head. The arm comprises a C-shaped
`“elbow” member, pivotally attached to a “shoulder”
`member. In turn, the “shoulder” member of the arm is
`pivotally connected to the housing. Dynamic couplings
`join the optical ?bers such that the shapes thereof re
`main ?xed regardless of the orientation of the arm.
`The housing further incorporates a Cartesian-coordi
`nate table for positioning the scanning head with re
`spect to a microplate that contains analyte samples. To
`measure absorbance, ?uorescence, and luminescence of
`30
`the samples, an optical system, incorporating a plurality
`of lenses, ?lters, and sensors is utilized. Radiant energy
`for these measurements is provided by a light source
`having a microcomputer-controlled power supply. The
`same microcomputer governs the operation of the opti
`35
`cal system and the positioning table.
`
`25
`
`45
`
`55
`
`4
`DETAILED DESCRIPTION
`Throughout the following description, speci?c de
`tails, such as materials, dimensions, etc., are set forth in
`order to provide a more thorough understanding of the
`invention. However, the invention may be practiced
`without these particulars. In other instances, well
`known elements have not been shown or described to
`avoid unnecessarily obscuring the present invention.
`Accordingly, the speci?cation and drawings are to be
`regarded in an illustrative, rather than a restrictive,
`sense.
`FIG. 1 shows a schematic side view of a multi-func
`tional photometer according to the present invention.
`The photometer comprises a housing 10 that pivotally
`supports a movable arm 12, containing a C-shaped rigid
`“elbow”member 14 and a rigid “shoulder” member 16.
`The housing is approximately 21 cm tall, 18 cm wide,
`and 26 cm long. Arm 12 incorporates a plurality of
`optical ?bers and is coupled to a ?rst scanning element,
`e.g., an optical scanning head 18. The structure of arm
`12 and the coupling mechanism of the optical ?bers will
`- be described fully in the ensuing section of the speci?ca
`tion.
`Scanning head 18 is rotationally attached through
`bearings 136 and 138 to a conventional positioning table
`20, e. g., the Pen Plotter table, manufactured by Hewlett
`Packard Company of Palo Alto, Calif. Positioning ta
`bles like the Pen Plotter are often computer controlled
`such that the computer speci?es X and Y coordinates of
`a point to be located by the mechanism of the table.
`Table 20 positions head 18 with respect to a microplate
`22 that holds samples to be analyzed in a multiplicity of
`analyte wells, such as a well 23. As illustrated in FIG. 1,
`both table 20 and microplate 22 are supported within
`housing 10.
`The optical system of the apparatus, described in
`reference to FIGS. 1 and 2, has a light-delivering assem
`bly, a light-gathering assembly for absorbance measure
`ments, and a light-gathering assembly for ?uorescence
`and luminescence measurements. The light-delivering
`assembly includes a light source 24; a collimating lens
`26; a plurality of bandpass ?lters 28, individually select
`able by means of a rotary ?lter wheel 30; a beam splitter
`32; a focusing lens 34; optical ?bers 36, 38, and 40 ar
`ranged in series; and a collimating lens 42. Light source
`24 typically comprises a xenon arc lamp, energized by a
`DC power supply 44, e.g., of Type 5 manufactured by
`Mimir Corporation of Sunnyvale, Calif. The power
`supply is controlled by a microcomputer 46, which also
`governs the positioning operations of table 20 and the
`functions of the optical system, e. g., the angular position
`of ?lter wheel 30. Microcomputer 46 may have, for
`example, a 80286 microprocessor from Intel Corpora
`tion of Santa Clara, Calif.
`The light-gathering assembly for absorbance mea
`surements comprises a reference-signal photodetector
`48, a focusing lens 50, and a second scanning element
`for collecting light transmitted through microplate 22,
`e.g., a photodetector 52. Photodetectors 48 and 52,
`which convert electromagnetic radiation into electric
`current, may be implemented as photovoltaic cells.
`After being converted to a digital format by an analog
`to-digital converter (not shown), the outputs of photo
`detectors 48 and 52 are analyzed by microcomputer 46.
`The light-gathering assembly for ?uorescence and
`luminescence measurements includes optical pick-up
`?bers 54, 56, and 58, arranged side-by-side. The pick-up
`
`BRIEF DESCRIPTION OF THE DRAWINGS
`The present invention is illustrated by way of exam
`ple, and not by way of limitation, in the ?gures of the
`accompanying drawings, where:
`FIG. 1 is a schematic side view of a multi-functional
`photometer according to the present invention.
`FIG. 2 is a schematic representation of an optical
`system utilized by the photometer of FIG. 1.
`FIG. 3 is a schematic representation of an alternative
`embodiment of the optical system of FIG. 2.
`FIG. 4 is a side elevational view of a movable arm of
`the photometer illustrated in FIG. 1.
`FIG. 5 is a sectional view of an optical-?ber coupler
`of the photometer of FIG. 1.
`FIG. 5A is a sectional view of an optical-?ber cou
`pling provided by the couplers such as the one shown in
`FIG. 5.
`FIG. 6 is a sectional view of an optical scanning head
`of the photometer shown in FIG. 1.
`FIG. 7 is a top plan view of the scanning head of
`FIG. 6.
`FIG. 8 is a sectional view of an optical-?ber integra
`tor utilized by the photometer of FIG. 1.
`FIG. 9 is a side elevational view of the movable arm
`of FIG. 4 modi?ed to accommodate the alternative
`embodiment of the optical system, shown in FIG. 3.
`FIG. 10 is a block diagram illustrating the operation
`of the photometer of FIG. 1.
`For purposes of illustration, these ?gures are not
`necessarily drawn to scale. In all of the ?gures, like
`components are designated by like reference numerals.
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`?bers are collectively coupled to a light-transmitting
`Member 14 further includes parallel beams 122 and
`?ber 60, which interfaces with an optical ?ber 62. Upon
`124, integrally connected by a shank 126. Beam 122
`exiting ?ber 62, light passes through a collimating lens
`contains a cylindrical bore 128 that accommodates
`64; one of a plurality of bandpass ?lters 66, selectable by
`scanning head 18 (?rst scanning element) whereas beam
`turning a rotary ?lter wheel 68, which is computer-con
`124 bears the second scanning element comprising lens
`50 and photodetector 52. The second scanning element,
`trolled; and a focusing lens 70. Lens 70 focuses the
`optical signal on a photodetector 72, whose output is
`which is collinear with the scanning head, is located
`then digitized and processed by microcomputer 46. In
`with respect to beam 124 with dowel pins (not shown)
`an alternative embodiment of the optical system (FIG.
`and is attached to the beam with screw-type fasteners.
`Head 18 comprises a substantially cylindrical casing
`3), a light-dispersing device 74 replaces ?lter wheel 68
`for ?uorescence and luminescence measurements.
`130 that is retained inside bore 128, e. g., with a set screw
`Moreover, instead of being directed to photodetector
`131. Casing 130 has a through longitudinal opening 132
`52, the optical signal, transmitted through one of a mul
`that houses an optical-?ber coupler 134 at one end and
`tiplicity of microplate wells 23, is channeled to the
`lens 42 at the other. A set screw 135 anchors coupler
`light-dispersing device through lens 50 via sequentially
`134 within opening 132. Ring bearings 136 and 138 are
`coupled optical ?bers 76, 78, and 80. Light-dispersing
`mounted on ?anges de?ning bore 128 for rotationally
`device 74 comprises a diffraction grating that disperses
`coupling head 18 to positioning table 20 (schematically
`incoming optical radiation into component wave
`shown in FIG. 1). Casing 130 further comprises three
`lengths, which are gathered at photodetector 72. Thus,
`through cavities 140 (only one of which is shown in
`analyses of optical signals resulting from phenomena of
`FIG. 4), symmetrically arranged around opening 132
`and having an angle of approximately 12° with respect
`absorbance, ?uorescence, and luminescence can be per
`to the vertical axis of the casing.
`formed over a range of wavelengths, rather than at a
`narrow spectral bandwidth provided by an individual
`Cavities 140 contain ends of optical ?bers 54, 56, and
`?lter. Consequently, valuable additional information
`58, which may be used to pick up fluorescent emissions.
`may be learned about the properties of analyte samples
`Due to the oblique arrangement of cavities 140, these
`being studied.
`?bers are less likely to receive excitation from ?ber 40.
`The opposite ends of ?bers 54, 56, and 58 are routed via
`a lateral opening in sleeve 118 into an optical-?ber inte
`grator 142, which contains a through central opening
`for housing the ?bers. Integrator 142 is anchored by a
`set screw 146 inside a through central bore of spacer
`148, the latter being ?xed by the same screw within
`sleeve 118. The integrator is positioned such that its
`central opening is collinear with the central bore of
`coupler 96 to allow exchange of radiant energy between
`?ber 60 and ?bers 54, 56, and 56.
`A set screw 150 secures an optical-?ber coupler 152,
`identical to couplers 90, 92, 94, 96, and 134, within a
`through opening in hinge portion 116 such that the
`bores of couplers 94 and 152 are collinear. The above
`described couplers may be made of an opaque material,
`such as aluminum. Each coupler is about 6.1 mm long
`and the radial dimension of the longitudinal central bore
`is approximately 0.5 mm. The optical ?bers inserted
`inside the couplers, e.g., couplers 94 and 152, com
`pletely occupy central bores 99 such that the ends of the
`?bers are ?ush with the end-faces of the couplers, as
`shown in FIG. 5A. The ?bers are typically retained
`inside the couplers by friction or with an adhesive
`placed along the ?ber shafts such that during insertion
`of the ?bers into the couplers the end-faces of the ?bers
`are not covered with the adhesive.
`Casing 130 of scanning head 18 is illustrated in
`greater detail in FIGS. 6 and 7. A sectional view of the
`casing (FIG. 6) depicts the con?guration of opening
`132, which comprises a coupler portion 154 and a lens
`portion 156. Portion 154 houses coupler 134 (shown in
`FIG. 4) while portion 156 is used for mounting collimat
`ing lens 42 (shown in FIGS. 1 and 4). The path of radi
`ant energy through the casing is restricted by a neck
`aperture 158 formed in casing 130. Inclined, through
`cavities 140, only one of which can be shown in the
`sectional view of FIG. 6, surround opening 132. The
`cavities contain optical ?bers, such as ?ber 54, and are
`equidistant from each other (FIG. 7). The ?bers occupy
`the full length of cavities 140 such that the ends of the
`?bers are flush or only slightly recessed with respect to
`the endface of casing 130. The casing may be made of an
`
`MOVABLE ARM FOR ROUTING OPTICAL
`FIBERS
`Movable arm 12, generally illustrated in FIG. 1, is
`shown in greater detail in FIG. 4. The arm comprises an
`articulated linkage having movably coupled members
`14 and 16. Member 16 is a substantially rectangular
`structure having mounting protrusions 82, 84, 86, and
`88. The protrusions contain openings accommodating
`optical-?ber couplers 90, 92, 94, and 96, respectively.
`The couplers are ?xed inside the openings with
`threaded fasteners, e.g., set screws 98.
`As shown in FIG. 5, each of the couplers of the type
`described above, for example coupler 96, contains a
`centrally-disposed through bore 99, having a radial
`dimension that is uniform along the entire length of the
`bore. Moreover, each coupler has two distinct cylindri
`cal surfaces 101 and 103. Surface 101 has a larger radial
`dimension then surface 103 and de?nes the end of the
`coupler where an optical ?ber is to be inserted.
`FIG. 4 further illustrates the pivotal attachment of
`member 16 to housing 10 by means of a bearing assem
`bly 100, which includes a pair of ring bearings 102 and
`104 that support couplers 90 and 92. Bearings 102 and
`104 are retained within collars 106 and 108, respec
`tively, where collar 108 is integral with housing 10. The
`two collars are rigidly interconnected by a hollow cy
`lindrical sleeve 110. The above-described structure
`allows member 16 to pivot with respect to housing 10
`55
`about an axis de?ned by the vertical symmetry axis of
`sleeve 110.
`Similarly, bearings 112 and 114 allow member 16 to
`pivotally support C-shaped member 14. The C-shaped
`member has a hinge portion 116, which is rigidly at
`tached to one end of a cylindrical hollow sleeve 118
`with a set screw 120. The inner races of bearings 112
`and 114 are mounted on couplers 96 and 94, respec
`tively. The outer race of bearing 114 sustains portion
`116, while bearing 112 is inserted into the second end of
`65
`sleeve 118. This structure permits member 14 to pivot
`with respect to member 16 about an axis de?ned by the
`vertical symmetry axis of sleeve 118.
`
`35
`
`45
`
`THERMO FISHER EX. 1031
`
`

`
`8
`DYNAMIC OPTICAL-FIBER COUPLING
`PROVIDED BY MOVABLE ARM
`The operation of dynamic optical-?ber couplings
`provided by arm 12 can now be outlined with reference
`to FIGS. 4 and 5A.
`As table 20 positions scanning head 18 at various
`wells of the microplate, member 14 pivots on bearings
`112 and 114 relative to member 16. In turn, member 16
`pivots relative to housing 10 on bearings 102 and 104.
`Speci?cally, as member 14 rotates with respect to mem
`ber 16, ?bers 40, 54, 56, and 58 move together therewith
`without twisting or bending. Optical contact between
`?ber 40 and ?ber 38 is maintained through the dynamic
`coupling provided by couplers 152 and 94 regardless of
`the angular relationship between members 14 and 16.
`Optical contact between ?ber 60 and pick-up ?bers 54,
`56, and 58 is maintained in a similar manner with the use
`of coupler 96 and integrator 142. Moreover, the integra
`tor allows the system to relay the optical signals of a
`plurality of ?bers into a single ?ber, thus providing a
`simple, yet extremely sensitive optical arrangement for
`performing ?uorescence measurements.
`Fibers 38 and 60 are also dynamically coupled with
`?bers 36 and 62, respectively, since couplers 90 and 92
`rotate relative to housing 10 in respective bearings 102
`and 104, whereas ?bers 36 and 62 remain stationary in
`couplers 109 and 107, which are anchored to collars 106
`and 108 of housing 10.
`Thus, bending and twisting of optical ?bers is elimi
`nated, guaranteeing repeatability and consistency of
`measurements and preventing mechanical failure of
`?ber cores due to cyclical bending stresses. Moreover,
`since compliance of optical ?bers does not affect the
`movement of scanning head 18, stiffer quartz ?bers can
`now be employed to allow transmission of ultraviolet
`radiation, which may be useful in certain types of ?uo
`rescence measurements. Also, the absence of bending
`resistance in the ?bers permits the positioning table to
`move the scanning head quickly enough to perform
`kinetic measurements.
`Additionally, parallel beams 122 and 124 of member
`14 allow the system to position lens 50 and photodetec
`tor 52 collinearly with respect to scanning head 18 so
`that absorbance measurements (typically done by pass
`ing radiant energy from ?ber 40-to detector 52 through
`an analyte sample) can be performed together with
`?uorescence and luminescence assays. Furthermore,
`the scanning head orients the ends of optical ?bers 54,
`56, and 58 obliquely to its longitudinal axis to prevent
`the ?bers from picking up optical noise from the edges
`of microplate wells during fluorescence and lumines
`cence measurements. Fibers 54, 56, and 58 are designed
`to pick up (receive) ?ourescence and luminescence
`emissions and ?ber 40 is designated to provide the exci
`tation light in the case of ?uorescence. In this manner,
`?uorescence and luminescence measurements are taken
`above the microplate rather than through it.
`
`5,436,718
`7
`opaque material, e.g., aluminum, and is approximately
`17.5 mm long. Neck aperture 158 restricts the diameter
`of the light path to approximately 2.0 mm.
`The construction of optical-?ber integrator 142 is
`described in detail with reference to FIG. 8. Integrator
`142 has a generally cylindrical shape and a centrally-dis
`posed aperture 144 containing an optical ?ber segment
`159 that is secured inside the aperture, e.g., by an adhe
`sive. Segment 159 completely ?lls aperture 144 such
`10
`that one end of the segment is flush with the endface of
`integrator 142. The integrator also possesses a cylindri
`cal bore 160 that accommodates the ends of optical
`?bers 54, 56, and 58, which are ?xed inside the bore.
`Bore 160 has a slightly greater radius than aperture 144
`and is joined therewith at a ?ange 162 so that ?ber
`segment 159 is contiguous with ?bers 54, 56, and 58. To
`maximize light transmission between ?ber segment 159
`and ?bers 54, 56, and 58, the difference in the radial
`dimensions of aperture 144 and bore 160 is minimal.
`Thus, ?bers 54, 56, and 58 each have a smaller diameter
`than segment 159 such that their combined cross-sec
`tional area is approximately the same as that of segment
`159. In turn, the diameter of segment 159 is the same as
`those of ?bers 36, 38, 40, 60, and 62. To facilitate the
`insertion of the optical ?bers into the integrator, an
`opening 164, which possesses a greater radius than the
`bore, is formed collinearly with the latter. Opening 164
`has a countersink 165 for gradually guiding the ends of
`the optical ?bers into bore 160. Integrator 142 may be
`made of an opaque material, e.g., aluminum. The inte
`grator is about 25 mm long, aperture 144 is about 2.3
`mm in diameter, and bore 160 has a diameter of approxi
`mately 2.3 mm.
`Referring once again to FIG. 4, the coupling of the
`optical ?bers is now further described. One end of ?ber
`40 is secured inside the bore of coupler 134 while the
`other end is routed inside coupler 152 via a lateral open
`ing within sleeve 118. Similarly, the ends of ?ber 38 are
`retained inside couplers 92 and 94. Fiber 60 and cou
`40
`plers 90 and 96 are arranged identically. A space of
`about 0.2 mm is provided between the juxtaposed faces
`of couplers 94 and 152 as well as between those of cou
`pler 96 and integrator 142 for allowing member 14 of
`the movable arm to freely pivot with respect to member
`45
`16.
`To link ?bers 38 and 60 with the optical system de
`scribed in the previous section of the speci?cation, opti
`cal ?bers 36 and 62, interfacing with the rest of the
`optical components, are routed into sleeve 110 via a
`lateral opening therein. The ends of these ?bers are
`supported within couplers 107 and 109 anchored inside
`collars 106 and 108 such that the ?bers 36 and 62 are
`collinear with ?bers 38 and 60, respectively. A distance
`of approximately 0.2 mm separates the contiguous faces
`of couplers 90 and 107 as well as the faces of couplers 92
`and 109. This permits member 16 to pivot freely with
`respect to housing 10. Fibers 38, 40, and 60 are approxi
`mately 20 cm long and 1.0 mm in diameter. Fibers 54,
`56, and 58 each have a diameter of about 0.8 mm and a
`length of approximately 20 cm. In one embodiment of
`the invention, all optical ?bers are made of quartz,
`thereby allowing transmission of ultraviolet light.
`As noted above, the juxtaposed faces of the respec
`tive couplers (e.g., 94 and 152) are aligned such that the
`ends of their respective ?bers are collinear and contigu
`ous to maximize light transmission between the ?bers.
`The alingment of the ?bers is illustrated in FIG. 5A.
`
`65
`
`MOVABLE ARM MODIFIED TO
`ACCOMMODATE ALTERNATIVE
`EMBODIMENT OF OPTICAL SYSTEM
`
`FIG. 9 shows a movable arm modi?ed to accommo
`date the alternative embodiment‘of the optical system
`(illustrated in FIG. 3).
`In order to provide an optical connection between
`the second scanning