United States Patent [19J
`Fernandes et al.
`
`[54] MUTLI-FUNCTIONAL PHOTOMETER
`WITH MOVABLE LINKAGE FOR ROUTING
`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.6 ....•.......••.....•• GOIN 21/59; GOIN 21/64;
`G02B 6/24
`[52] U.S. Cl. ...................................... 356173; 356/318;
`356/417; 356/418; 356/436; 356/440;
`250/458.1; 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
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`3,697.185 10/1972 Kassel et al. ........................ 356/410
`3,874,780 4/1975 Love .
`4.373.779 2/1983 Dorsey .
`4,477, 190 10/1984 Liston et al. ........................ 356/418
`4,501,970 2/1985 Nelson .............................. 250/458.1
`4,587,812 5/1986 Brega .
`4,622,468 11/1986 Stefanski et al. ................. 250/458.1
`4.626,684 12/1986 Landa .................................. 356/318
`4,669,878 6/1987 Meier .................................. 356/319
`4,730,922 3/1988 Bach et al. ............................ 356/73
`4, 750,837 6/1988 Gifford et al. ...................... 356/417
`4,799,756 1/1989 Hirschfeld ........................ 250/458.1
`4,802,768 2/1989 Gifford et al. ...................... 356/417
`4,815,812 3/1989 Miller .
`4,820,045 4/1989 Boisde et al. ....................... 356/319
`
`I lllll llllllll Ill lllll lllll lllll lllll lllll 111111111111111111111111111111111
`5,436, 718
`Jul. 25, 1995
`
`US005436718A
`[11] Patent Number:
`[45] Date of Patent:
`
`4,840,485 6/1989 Gratton ............................... 356/317
`4,937,457 6/1990 Mitchell ........................... 250/458.1
`4,945,245 7/1990 Levin ............................... 250/458.1
`4.968,148 11/1990 Chow et al. ........................ 356/427
`5,030,832 7 /1991 Williams et al. ................. 250/458.1
`5.125,747 6/1992 Sayegh et al. ...................... 356/407
`5.131,746 7/1992 O'Rourke et al. .................. 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 .............................. 250/458.l
`
`FOREIGN PATENT DOCUMENTS
`
`62160 10/1982 European Pat. Off. ............ 356/414
`
`OTHER PUB LI CA TIO NS
`''The Cytoflour Flourescence Measurement System".
`Automated Flourescence Scanning, Millipore, 1990.
`Primary Examiner-F. L. Evans
`Attorney, Agent, or Firm-Blakely, Sokoloff, Taylor &
`Zafman
`
`[57]
`ABSTRACT
`A multi-functional photometer includes a scanning
`mechanism having a housing (10) that bears a movable
`linkage (12). The linkage is coupled to an optical scan(cid:173)
`ning head (18) and incorporates optical fibers for trans(cid:173)
`mitting radiant energy to and from the scanning head.
`The arm comprises a C-shaped "elbow" member (14),
`pivotally attached to a "shoulder" member (16). In tum,
`the "shoulder" member of the arm is pivotally con(cid:173)
`nected to the housing. Dynamic couplings join the opti(cid:173)
`cal fibers such th-at the shapes thereof remain fixed re(cid:173)
`gardless of the orientation of the arm. The housing
`further incorporates a Cartesian-coordinate table (20)
`for positioning the scanning head with respect to a
`microplate (22) that contains a plurality of analyte sam(cid:173)
`ples.
`
`47 Claims, 10 Drawing Sheets
`
`48
`
`12
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 1of10
`
`5,436,718
`
`0
`
`CD
`¢
`
`..
`
`- •
`(!) -IJ..
`
`"C\J
`
`co/
`f'()
`
`¢
`f'()
`
`~ It\ CD
`~ .
`~ ~m
`~
`0
`,.....
`
`C\J
`r<:I
`
`Cl)
`C\J
`
`0
`rQ
`
`CD
`C\J
`
`a:>(cid:173)w...J
`~!5 _ __.
`,c..
`
`Cl.(/)
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 2 of 10
`
`5,436,718
`
`N .
`(!)
`LL
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 3 of 10
`
`. 5,436,718
`
`r<> .
`
`(.!)
`LL
`
`THERMO FISHER EX. 1022
`
`

`
`QO
`~
`
`~ .., ._..
`~ w
`..,
`C.11
`
`0
`~
`...+
`('!) ro
`l:r"
`'J1
`
`'""' >"""
`
`0
`
`62
`
`01
`~
`~
`>"""
`
`;;--~
`
`_,01
`N
`
`~ (D = f"+-
`
`l"tJ
`•
`00
`c •
`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 4 of 10
`
`5,436,718
`
`W
`
`/////////
`u///41
`
`6)
`fl‘
`®
`
`7' -
`
`60
`
`14
`/
`
`124
`
`126
`
`\..
`
`12
`
`\
`
`I-1 [.\\\\\\\\\ -
`
`I
`5-II\\‘.\'x\\\\‘
`I
`
`FIG.4
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 5 of 10
`
`5,436,718
`
`101
`
`96
`
`/
`
`Fl G.5
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 6 of 10
`
`5,436,718
`
`co
`
`(\j
`LO
`
`0 v
`
`<(
`IO .
`(!)
`LL
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 7 of 10
`
`5,436,718
`
`132
`t
`
`154
`
`140
`
`156
`FIG. 6
`
`154
`
`140
`
`FIG.7
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 8of10
`
`5,436,718
`
`142
`
`FI G.8
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 9 of 10
`
`5,436,718
`
`0 ~
`
`CX)
`
`0
`<O
`
`'
`
`<O
`
`•
`
`CX)
`
`"' m
`I") (.!) -LL
`
`THERMO FISHER EX. 1022
`
`

`
`U.S. Patent
`
`July 25, 1995
`
`Sheet 10 of 10
`
`5,436,718
`
`START
`
`200
`
`IDLE LIGHT 1-------1---(cid:173)
`SOURCE
`
`,.-----'---- 202
`
`NO
`
`OPERATING
`INSTRUCTIONS
`RECEIVED?
`
`YES
`
`SELECT
`SPECIFIED
`BANDPASS
`FILTERS
`
`POWER-UP
`LIGHT
`SOURCE
`
`204
`
`206
`
`208
`
`NO
`
`MOVE
`SCANNING
`HEAD 11HOME11
`AND CALIBRATE
`PHOTODETECTORS
`
`210
`
`YES
`
`FI G.10
`
`THERMO FISHER EX. 1022
`
`

`
`1
`
`5,436,718
`
`2
`croplates are beneficial since they allow simultaneous
`preparation of a large number of test samples. More(cid:173)
`over, microplates are inexpensive, safe, sturdy, and
`convenient to handle. They are also disposable and can
`5 be cleaned easily when necessary.
`One instrument currently available for fluorescent
`analysis of samples in microplate wells is the Cytofluor
`2300 fluorometer, distributed by Millipore Corporation,
`Bedford, Mass. This fluorometer includes a scanning
`10 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 fibers that transmits
`excitation and emission radiation.
`However, the capabilities of the Cytofluor 2300 flu(cid:173)
`orometer are limited in that it cannot perform absor(cid:173)
`bance measurements. Furthermore, the movement of
`the scanning head from one microplate well to another
`continuously alters the geometrical configuration of the
`optical-fiber bundle that is attached to the head. Conse(cid:173)
`quently, curvatures of the light-transmitting fibers
`change, introducing variations in their optical proper(cid:173)
`ties. These variations create inconsistencies in readings
`between different wells and adversely affect the repeat(cid:173)
`ability, and thus, accuracy of measurements. Moreover,
`continuous bending of the fibers produces stresses that
`cause mechanical failure of the fiber cores.
`Additionally, to allow unrestricted movement of the
`scanning head, flexible plastic fibers are employed, as
`opposed to less pliable quartz fibers. On the down side,
`plastic fibers cannot efficiently transmit radiant energy
`in the ultraviolet (UV) region of the spectrum. Accord(cid:173)
`ingly, the fluorometer is unable to perform measure-
`ments, such as binding studies of certain proteins, e.g.,
`tryptophan, since fluorescence analyses of this type
`require the use of UV radiation. Furthermore, the de(cid:173)
`formation resistance of the optical-fiber bundle slows
`the movements of the scanning head, thus limiting the
`ability of the apparatus to perform kinetic measure(cid:173)
`ments.
`Another spectroscopic apparatus utilizing micro-
`plates is disclosed in U.S. Pat. No. 4,968,148 to Chow et
`al., 1990. Chow's device uses an optical distributing
`element to selectively direct radiant energy to specified
`microplate sites. One drawback of this instrument is its
`inability to perform fluorescence measurements. More(cid:173)
`over, the large number of fibers unnecessarily compli(cid:173)
`cates the apparatus and increases production costs.
`Also, the light-delivery system of the instrument has a
`fixed 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(cid:173)
`plates having different configurations of wells.
`
`MUTLI-FUNCTIONAL PHOTOMETER WITH
`MOY ABLE LINKAGE FOR ROUTING OPTICAL
`FIBERS
`
`FIELD OF THE INVENTION
`The present invention relates to the field of spectros(cid:173)
`copy, particularly to a multi-functional photometer
`capable of measuring light absorbance, fluorescence,
`and luminescence of a sample.
`
`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 15
`and interpretation of electromagnetic radiation ab(cid:173)
`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- 20
`ments of absorbance, fluorescence, and luminescence.
`Chemical analyses with absorption spectroscopy
`allow one to determine concentrations of specific com(cid:173)
`ponents, to assay chemical reactions, and to identify
`individual compounds. Absorbance measurements are 25
`most commonly used to find the concentration of a
`specific 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- 30
`ple.
`Fluorescence, in turn, is a physical phenomenon
`based upon the ability of some substances to absorb and
`subsequently emit electromagnetic radiation. The emit(cid:173)
`ted radiation has a lower_ energy level and a longer 35
`wavelength than the excitation radiation. Moreover, the
`absorption of light is wavelength dependent. In other
`words, a fluorescent substance emits light only when
`the excitation radiation is in the particular excitation
`band (or bands) of that substance.
`For fluorescence measurements, fluorescent dyes
`called fluorophores are often used to "tag" molecules of
`interest, or targets. After being irradiated by an excita(cid:173)
`tion beam, fluorophores, bonded to the targets, emit
`light that is then collected and quantized. The ratio of 45
`the intensity of the emitted fluorescent light to the in(cid:173)
`tensity of the excitation light is called the "relative
`fluorescence intensity" and serves as an indicator of
`target concentration. Another useful characteristic is
`the phase relationship between the cyclic variations in 50
`the emitted light and the variations in the excitation
`light, i.e., the time lag between corresponding varia(cid:173)
`tions in the emission and excitation beams.
`As noted above, luminescence measurements can also
`be employed for analyzing biological samples. Lumines- 55
`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(cid:173)
`cence can-be produced by energy-transfer mechanisms
`that take energy of a high intensity, e.g., a radioactive 60
`emission, and transform it to energy of a low intensity,
`e.g., a flash of light.
`At the present time, a variety of spectroscopic instru(cid:173)
`ments is commonly used in the art. A number of these
`instruments are designed to be utilized in conjunction 65
`with multi-site analyte receptacles called "microplates'',
`which usually comprise one-piece structures having
`multiplicities of wells for holding analyte samples. Mi-
`
`40
`
`OBJECTS AND SUMMARY OF THE
`INVENTION
`It is accordingly an object of the invention to provide
`a multi~functional photometer which overcomes the
`foregoing disadvantages, e.g., which measures absor(cid:173)
`bance, fluorescence, and luminescence of a sample;
`which provides repeatable measurements and produces
`consistent readings between different test sites; which
`eliminates recurring bending of optical fibers and me(cid:173)
`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-
`
`THERMO FISHER EX. 1022
`
`

`
`5,436,718
`
`3
`figurations; and which is relatively simple and inexpen(cid:173)
`sive to manufacture.
`Another object of the invention is to supply a pho(cid:173)
`tometer having a movable linkage for dynamically and
`interconnectingly routing optical fibers such that a con- 5
`stant configuration 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, fluorescence, 10
`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, 15
`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 fibers, for transmitting radiant energy to and 20
`from the scanning head. The arm comprises a C-shaped
`"elbow" member, pivotally attached to a "shoulder"
`member. In tum, the "shoulder" member of the arm is
`pivotally connected to the housing. Dynamic couplings
`join the optical fibers such that the shapes thereof re- 25
`main fixed regardless of the orientation of the arm.
`The housing further incorporates a Cartesian-coordi(cid:173)
`nate table for positioning the scanning head with re(cid:173)
`spect to a microplate that contains analyte samples. To
`measure absorbance, fluorescence, and luminescence of 30
`the samples, an optical system, incorporating a plurality
`of lenses, filters, 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.
`
`4
`DETAILED DESCRIPTION
`Throughout the following description, specific de(cid:173)
`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 specification 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(cid:173)
`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 fibers and is coupled to a first scanning element,
`e.g., an optical scanning head 18. The structure of arm
`12 and the coupling mechanism of the optical fibers will
`· be described fully in the ensuing section of the specifica(cid:173)
`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(cid:173)
`bles like the Pen Plotter are often computer controlled
`such that the computer specifies 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
`BRIEF DESCRIPTION OF THE DRAWINGS
`reference to FIGS. 1and2, has a light-delivering assem-
`bly, a light-gathering assembly for absorbance measure-
`The present invention is illustrated by way of exam-
`ple, and not by way of limitation, in the figures of the 40 ments, and a light-gathering assembly for fluorescence
`accompanying drawings, where:
`and luminescence measurements. The light-delivering
`FIG. 1 is a schematic side view of a multi-functional
`assembly includes a light source 24; a collimating lens
`photometer according to the present invention.
`26; a plurality of bandpass filters 28, individually select-
`FIG. 2 is a schematic representation of an optical
`able by means of a rotary filter wheel 30; a beam splitter
`system utilized by the photometer of FIG. 1.
`45 32; a focusing lens 34; optical fibers 36, 38, and 40 ar-
`FIG. 3 is a schematic representation of an alternative
`ranged in series; and a collimating lens 42. Light source
`embodiment of the optical system of FIG. 2.
`24 typically comprises a xenon arc lamp, energized by a
`FIG. 4 is a side elevational view of a movable arm of DC power supply 44, e.g., of Type 5 manufactured by
`the photometer illustrated in FIG. 1.
`Mimir Corporation of Sunnyvale, Calif. The power
`FIG. 5 is a sectional view of an optical-fiber coupler 50 supply is controlled by a microcomputer 46, which also
`of the photometer of FIG. 1.
`governs the positioning operations of table 20 and the
`FIG. SA is a sectional view of an optical-fiber cou-
`functions of the optical system, e.g., the angular position
`piing provided by the couplers such as the one shown in
`of filter wheel 30. Microcomputer 46 may have, for
`FIG. 5.
`example, a 80286 microprocessor from Intel Corpora-
`FIG. 6 is a sectional view of an optical scanning head 55 tion of Santa Clara, Calif.
`of the photometer shown in FIG. 1.
`The light-gathering assembly for absorbance mea-
`FIG. 7 is a top plan view of the scanning head of
`surements comprises a reference-signal photodetector
`FIG. 6.
`48, a focusing lens 50, and a second scanning element
`FIG. 8 is a sectional view of an optical-fiber integra-
`for collecting light transmitted through microplate 22,
`tor utilized by the photometer of FIG. 1.
`60 e.g., a photodetector 52. Photodetectors 48 and 52,
`FIG. 9 is a side elevational view of the movable arm
`which convert electromagnetic radiation into electric
`of FIG. 4 modified to accommodate the alternative
`current, may be implemented as photovoltaic cells.
`embodiment of the optical system, shown in FIG. 3.
`After being converted to a digital format by an analog-
`FIG. 10 is a block diagram illustrating the operation
`to-digital converter (not shown), the outputs of photo-
`of the photometer of FIG. 1.
`65 detectors 48 and 52 are analyzed by microcomputer 46.
`For purposes of illustration, these figures are not
`The light-gathering assembly for fluorescence and
`necessarily drawn to scale. In all of the figures, like
`luminescence measurements includes optical pick-up
`components are designated by like reference numerals.
`fibers 54, 56, and 58, arranged side-by-side. The pick-up
`
`THERMO FISHER EX. 1022
`
`

`
`5
`fibers are collectively coupled to a light-transmitting
`fiber 60, which interfaces with an optical fiber 62. Upon
`exiting fiber 62, light passes through a collimating lens
`64; one of a plurality of bandpass filters 66, selectable by
`turning a rotary filter wheel 68, which is computer-con- 5
`trolled; and a focusing lens 70. Lens 70 focuses the
`optical signal on a photodetector 72, whose output is
`then digitized and processed by microcomputer 46. In
`an alternative embodiment of the optical system (FIG.
`3), a light-dispersing device 74 replaces filter wheel 68 10
`for fluorescence and
`luminescence measurements.
`Moreover, instead of being directed to photodetector
`52, the optical signal, transmitted through one of a mul(cid:173)
`tiplicity of microplate wells 23, is channeled to the
`light-dispersing device through lens 50 via sequentially 15
`coupled optical fibers 76, 78, and 80. Light-dispersing
`device 74 comprises a diffraction grating that disperses
`incoming optical radiation into component wave(cid:173)
`lengths, which are gathered at photodetector 72. Thus,
`analyses of optical signals resulting from phenomena of 20
`absorbance, fluorescence, and luminescence can be per(cid:173)
`formed over a range of wavelengths, rather than at a
`narrow spectral bandwidth provided by an individual
`filter. Consequently, valuable additional information
`may be learned about the properties of analyte samples 25
`being studied.
`
`MOVABLE ARM FOR ROUTING OPTICAL
`FIBERS
`Movable arm 12, generally illustrated in FIG. 1, is 30
`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 35
`optical-fiber couplers 90, 92, 94, and 96, respectively.
`The couplers are fixed
`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 40
`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(cid:173)
`cal surfaces 101 and 103. Surface 101 has a larger radial
`dimension then surface 103 and defines the end of the 45
`coupler where an optical fiber is to be inserted.
`FIG. 4 further illustrates the pivotal attachment of
`member 16 to housing 10 by means of a bearing assem(cid:173)
`bly 100, which includes a pair of ring bearings 102 and
`104 that support couplers 90 and 92. Bearings 102 and 50
`104 are retained within collars 106 and 108, respec(cid:173)
`tively, where collar 108 is integral with housing 10. The
`two collars are rigidly interconnected by a hollow cy(cid:173)
`lindrical sleeve 110. The above-described structure
`allows member 16 to pivot with respect to housing 10 55
`about an axis defined 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- 60
`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(cid:173)
`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 defined by the
`vertical symmetry axis of sleeve 118.
`
`5,436,718
`
`6
`Member 14 further includes parallel beams 122 and
`124, integrally connected by a shank 126. Beam 122
`contains a cylindrical bore 128 that accommodates
`scanning head 18 (first scanning element) whereas beam
`124 bears the second scanning element comprising lens
`50 and photodetector 52. The second scanning element,
`which is collinear with the scanning head, is located
`with respect to beam 124 with dowel pins (not shown)
`and is attached to the beam with screw-type fasteners.
`Head 18 comprises a substantially cylindrical casing
`130 that is retained inside bore 128, e.g., with a set screw
`131. Casing 130 has a through longitudinal opening 132
`that houses an optical-fiber coupler 134 at one end and
`lens 42 at the other. A set screw 135 anchors coupler
`134 within opening 132. Ring bearings 136 and 138 are
`mounted on flanges defining bore 128 for rotationally
`coupling head 18 to positioning table 20 (schematically
`shown in FIG. 1). Casing 130 further comprises three
`through cavities 140 (only one of which is shown in
`FIG. 4), symmetrically arranged around opening 132
`and having an angle of approximately 12° with respect
`to the vertical axis of the casing.
`Cavities 140 contain ends of optical fibers 54, 56, and
`58, which may be used to pick up fluorescent emissions.
`Due to the oblique arrangement of cavities 140, these
`fibers are less likely to receive excitation from fiber 40.
`The opposite ends of fibers 54, 56, and 58 are routed via
`a lateral opening in sleeve 118 into an optical-fiber inte(cid:173)
`grator 142, which contains a through central opening
`for housing the fibers. Integrator 142 is anchored by a
`set screw 146 inside a through central bore of spacer
`148, the latter being fixed 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
`fiber 60 and fibers 54, 56, and 56.
`A set screw 150 secures an optical-fiber 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(cid:173)
`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 fibers inserted
`inside the couplers, e.g., couplers 94 and 152, com(cid:173)
`pletely occupy central bores 99 such that the ends of the
`fibers are flush with the end-faces of the couplers, as
`shown in FIG. SA. The fibers are typically retained
`inside the couplers by friction or with an adhesive
`placed along the fiber shafts such that during insertion
`of the fibers into the couplers the end-faces of the fibers
`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 configuration 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(cid:173)
`ing lens 42 (shown in FIGS. 1 and 4). The path of radi(cid:173)
`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 fibers, such as fiber 54, and are
`equidistant from each other (FIG. 7). The fibers occupy
`the full length of cavities 140 such that the ends of the
`fibers are flush or only slightly recessed with respect to
`the endface of casing 130. The casing may be made of an
`
`THERMO FISHER EX. 1022
`
`

`
`5,436,718
`
`25
`
`7
`opaque material, e.g., aluminum, and is approximately
`17.5 mm long. Neck aperture 1S8 restricts the diameter
`of the light path to approximately 2.0 mm.
`The construction of optical-fiber integrator 142 is
`described in detail with reference to FIG. 8. Integrator 5
`142 has a generally cylindrical shape and a centrally-dis(cid:173)
`posed aperture 144 containing an optical fiber segment
`1S9 that is secured inside the aperture, e.g., by an adhe(cid:173)
`sive. Segment 1S9 completely fills aperture 144 such
`that one end of the segment is flush with the endface of 10
`integrator 142. The integrator also possesses a cylindri(cid:173)
`cal bore 160 that accommodates the ends of optical
`fibers S4, S6, and S8, which are fixed inside the bore.
`Bore 160 has a slightly greater radius than aperture 144
`and is joined therewith at a flange 162 so that fiber 15
`segment 1S9 is contiguous with fibers S4, S6, and S8. To
`maximize light transmission between fiber segment 1S9
`and fibers S4, S6, and 58, the difference in the radial
`dimensions of aperture 144 and bore 160 is minimal.
`Thus, fibers S4, S6, and S8 each have a smaller diameter 20
`than segment 1S9 such that their combined cross-sec(cid:173)
`tional area is approximately the same as that of segment
`1S9. In turn, the diameter of segment 159 is the same as
`those of fibers 36, 38, 40, 60, and 62. To facilitate the
`insertion of the optical fibers 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 fibers into bore 160. Integrator 142 may be 30
`made of an opaque material, e.g., aluminum. The inte(cid:173)
`grator is about 25 mm long, aperture 144 is about 2.3
`mm in diameter, and bore 160 has a diameter of approxi(cid:173)
`mately 2.3 mm.
`Referring once again to FIG. 4, the coupling of the 35
`optical fibers is now further described. One end of fiber
`40 is secured inside the bore of coupler 134 while the
`other end is routed inside coupler 1S2 via a lateral open(cid:173)
`ing within sleeve 118. Similarly, the ends of fiber 38 are
`retained inside couplers 92 and 94. Fiber 60 and cou- 40
`piers 90 and 96 are arranged identically. A space of
`about 0.2 mm is provided between the juxtaposed faces
`of couplers 94 and 1S2 as well as between those of cou(cid:173)
`pler 96 and integrator 142 for allowing member 14 of
`the movable arm to freely pivot with respect to member 45
`16.
`To link fibers 38 and 60 with the optical system de(cid:173)
`scribed in the previous section of the specification, opti(cid:173)
`cal fibers 36 and 62, interfacing with the rest of the
`optical components, are routed into sleeve 110 via a 50
`lateral opening therein. The ends of these fibers are
`supported within couplers 107 and 109 anchored inside
`collars 106 and 108 such that the fibers 36 and 62 are
`collinear with fibers 38 and 60, respectively. A distance
`of approximately 0.2 mm separates the contiguous faces 55
`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(cid:173)
`mately 20 cm long and 1.0 mm in diameter. Fibers S4,
`S6, and S8 each have a diameter of about 0.8 mm and a 60
`length of approximately 20 cm. In one embodiment of
`the invention, all optical fibers are made of quartz,
`thereby allowing transmission of ultraviolet light.
`As noted above, the juxtaposed faces of the respec(cid:173)
`tive couplers (e.g., 94 and 1S2) are aligned such that the 65
`ends of their respective fibers are collinear and contigu(cid:173)
`ous to maximize light transmission between the fibers.
`The alingment of the fibers is illustrated in FIG. SA.
`
`8
`DYNAMIC OPTICAL-FIBER COUPLING
`PROVIDED BY MOY ABLE ARM
`The operation of dynamic optical-fiber couplings
`provided by arm 12 can now be outlined with reference
`to FIGS. 4 and SA.
`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.
`Specifically, as member 14 rotates with respect to mem(cid:173)
`ber 16, fibers 40, 54, S6, and S8 move together therewith
`without twisting or bending. Optical contact between
`fiber 40 and fiber 38 is maintained through the dynamic
`coupling provided by couplers 1S2 and 94 regardless of
`the angular relationship between members 14 and 16.
`Optical contact between fiber 60 and pick-up fibers S4,
`S6, and S8 is maintained in a similar manner with the use
`of coupler 96 and integrator 142. Moreover, the integra(cid:173)
`tor allows the system to relay the optical signals of a
`plurality of fibers into a single fiber, thus providing a
`simple, yet extremely sensitive optical arrangement for
`performing fluorescence measurements.
`Fibers 38 and 60 are also dynamically coupled with
`fibers 36 and 62, respectively, since couplers 90 and 92
`rotate relative to housing 10 in respective bearings 102
`and 104, whereas fibers 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 fibers is elimi(cid:173)
`nated, guaranteeing repeatability and consistency of
`measurements and preventing mechanical failure of
`fiber cores due to cyclical bending stresses. Moreover,
`since compliance of optical fibers does not affect the
`movement of scanning head 18, stiffer quartz fibers can
`now be employed to allow transmission of ultraviolet
`radiation, which may be useful in certain types of fluo(cid:173)
`rescence measurements. Also, the absence of bending
`resistance in the fibers permits the positioning table to
`mo