U.S. patent application number 11/662468 was filed with the patent office on 2007-10-25 for instrumentation and method adapted for optical measurement of an amplified luminescent proximity homogeneous assay.
This patent application is currently assigned to WALLAC OY. Invention is credited to Petri Kivela.
Application Number | 20070247628 11/662468 |
Document ID | / |
Family ID | 34958709 |
Filed Date | 2007-10-25 |
United States Patent
Application |
20070247628 |
Kind Code |
A1 |
Kivela; Petri |
October 25, 2007 |
Instrumentation and Method Adapted For Optical Measurement of an
Amplified Luminescent Proximity Homogeneous Assay
Abstract
The present invention relates generally to the field of
biochemical laboratory instrumentation for different applications
of measuring properties of samples on microtitration plates and
corresponding sample supports. An optical measurement
instrumentation is provided, a sample is activated and the emission
is detected, wherein between the activation and detection phases of
measuring the sample, a shift is made in the relative position
between the sample and elements directing the activation radiation
to the sample as well as in the relative position between the
sample and the elements receiving the emission radiation from the
sample. This can be implemented e.g. by moving the sample assay
plate and/or a measuring head between the activation and emission
phases of a sample. The invention allows a simultaneous activation
of a first sample and detecting emission from a second sample thus
enhancing efficiency of the measurement.
Inventors: |
Kivela; Petri;
(Piispanristi, FI) |
Correspondence
Address: |
YOUNG & THOMPSON
745 SOUTH 23RD STREET
2ND FLOOR
ARLINGTON
VA
22202
US
|
Assignee: |
WALLAC OY
Turku
FI
FI-20101
|
Family ID: |
34958709 |
Appl. No.: |
11/662468 |
Filed: |
September 10, 2004 |
PCT Filed: |
September 10, 2004 |
PCT NO: |
PCT/FI04/00525 |
371 Date: |
May 14, 2007 |
Current U.S.
Class: |
356/432 |
Current CPC
Class: |
G01N 2021/6441 20130101;
G01N 2021/6471 20130101; G01N 2201/0446 20130101; G01N 21/76
20130101; G01N 2021/6482 20130101; G01N 2021/6484 20130101; G01N
21/13 20130101; G01N 21/253 20130101; G01N 21/6452 20130101; G01N
2201/0453 20130101; G01N 2201/0826 20130101; G01N 2021/6421
20130101; G01N 2201/103 20130101; G01N 21/6428 20130101 |
Class at
Publication: |
356/432 |
International
Class: |
G01N 21/00 20060101
G01N021/00 |
Claims
1-18. (canceled)
19. An optical measurement instrument for measuring samples
(281-285), the optical measurement instrument comprising: a first
illumination source (211) disposed to produce first activation
radiation into a first sample (281), a first detector (231a)
disposed to measure first emission radiation from the first sample
(281) when the first sample is radiated with the first activation
radiation, shifting means (299) disposed to change a relative
position between the samples (281-285) and the first illumination
source and between the samples and the first detector,
characterized in that the optical measurement instrument further
comprises a second illumination source (212AS) disposed to produce
second activation radiation to a third sample (283), said shifting
means being disposed to change a relative position between the
samples (281-285) and the second illumination source.
20. An instrument according to claim 19, characterized in that the
first illumination source (211) and the first detector (231a) are
disposed to apply an Amplified Luminescent Proximity Homogeneous
Assay-analysing method.
21. An instrument according to claim 19, characterized in that said
first detector (231a) is disposed to measure photoluminescence
radiation.
22. An instrument according to claim 19, characterized in that said
second detector (291) is disposed to measure chemiluminescence
radiation.
23. An instrument according to claim 19, characterized in that said
first detector (231a) is disposed to measure both emission
radiation according to an Amplified Luminescent Proximity
Homogeneous Assay-analysis method and chemiluminescence
radiation.
24. An instrument according to claim 19, characterized in that the
second illumination source (212AS) is laser light source.
25. An instrument according to claim 19, characterized in that the
second detector (291) is one of the following: a photo-multiplier
tube and Charge Coupled Device.
26. An instrument according to claim 19, characterized in that the
first illumination source (211) and the second detector (291) are
adapted to operate simultaneously, the first illumination source
(211) producing the first activation radiation to the first sample
(281) and the second detector (291) measuring the second emission
radiation from the second sample (285).
27. An instrument according to claim 19, characterized in that the
shifting means (299) are disposed to change the relative position
between the samples and the first illumination source and between
the samples and the first detector by a distance that equals to a
distance between two adjacent samples or a multiple of said
distance between two adjacent samples.
28. An instrument according to claim 19, characterized in that the
shifting means (299) are disposed to change a relative position of
the first sample (285) and the second detector (291) in such a way
that the first emission radiation from the first sample is directed
to the second detector.
29. A method for optical measurement of samples, the method
comprising: directing (63) first activation radiation from a first
illumination source into a first sample, measuring (64) emission
radiation from a second sample, after the first sample has become
activated changing (66) a relative position between the samples and
the first illumination source in such a way that the first sample
is no more activated by the first activation radiation, measuring
(64) emission radiation from the first sample after the changing
(66) the relative position, characterized in that the method
further comprises directing second activating radiation from a
second illumination source into the first sample when measuring
(64) the emission radiation from the first sample after the
changing (66) the relative position.
30. A method according to claim 29, characterized in that the
measuring emission radiation comprises using an Amplified
Luminescent Proximity Homogeneous Assay-analysis method.
31. A method according to claim 29, characterized in that that the
measuring emission radiation comprises measuring
photoluminescence.
32. A method according to claim 29, characterized in that the first
activation radiation is laser light.
33. A method according to claim 29, characterized in that emission
radiation is measured using a photo-multiplier tube.
34. A method according to claim 29, characterized in that the
directing (63) the first activation radiation into the first sample
and the measuring (64) emission radiation from the second sample
are performed simultaneously.
35. A method according to claim 29, characterized in that a change
in the relative position equals to a distance between two adjacent
samples or a multiple of said distance between two adjacent
samples.
36. A method according to claim 29, characterized in that a first
detector is used for the measuring emission radiation from the
first sample when directing (63) the first activation radiation
into the first sample and a second detector is used for the
measuring (64) emission radiation from the first sample after the
changing (66) the relative position.
Description
BACKGROUND OF THE INVENTION
[0001] The present invention relates generally to the field of
biochemical laboratory instrumentation for different applications
of measuring properties of samples on e.g. microtitration plates
and corresponding sample supports. More particularly the invention
relates to more efficient, instrumental features of equipment used
as e.g. fluorometers, photometers and luminometers. The
applications may be e.g. clinical or research applications.
[0002] The routine work and also the research work in analytical
biochemical laboratories and in clinical laboratories is often
based on different tags or labels coupled on macromolecules under
inspection. The typical labels used are different radioactive
isotopes, enzymes, different fluorescent molecules and e.g.
fluorescent chelates of rare earth metals.
[0003] The detection of enzyme labels can be performed by utilizing
its natural biochemical function, i.e. to alter the physical
properties of molecules. In enzyme immunoassays colourless
substances are catalysed by enzyme into colourful substances or
non-fluorescent substances to fluorescent substances.
[0004] The colourful substances are measured with absorption, i.e.
photometric measurement. In the photometric measurement the
intensity of filtered and stabilized beam is first measured without
any sample and then the sample inside one plate is measured. The
absorbance i.e. the absorption values are then calculated.
[0005] The fluorescent measurement is generally used for measuring
quantities of fluorescent label substance in a sample. The most
photoluminescence labels are based on molecular photoluminescence
process. In this process optical radiation is absorbed by the
ground state of a molecule. Due to the absorption of energy the
quantum molecule rises into higher excited state. After the fast
vibrational relaxation the molecule returns back to its ground
state and the excess energy is released as an optical quantum. Due
to losses in this process the average absorbed energies are higher
than the average emitted energies. In the following, "activation"
is used as a term including excitation of photoluminescence as well
as other types of activation by radiation as is described
below.
[0006] A further measurement method is chemiluminescence
measurement where emission of a substance is measured from a sample
without activation by illumination. Thus a photoluminometer can
also be used as a chemiluminometer.
[0007] Further, there is an analysing method called Amplified
Luminescent Proximity Homogeneous Assay or AlphaScreen.TM.. The
function of the AlphaScreen method is based on the use of small
beads that attach to the molecules under study. There are two types
of beads that are coated with a material acting either as a donor
or acceptor of singlet-state oxygen. The measurement starts, when
the liquid sample is illuminated by light with wavelength of 680
nm. After this the material in the donor bead converts ambient
oxygen into singlet-state oxygen. The single-state molecules have a
short lifetime and they can reach only about a 200 nm distance by
diffusion in the liquid. If the chemical reaction in question has
taken place, both the donor and acceptor beads are bound to the
same molecule and so they are close to each other. In this case the
singlet-state oxygen may reach the acceptor bead where a series of
reactions is started. As the last phase of the reaction the coating
material in the acceptor beads emits photons in the 500-700 nm
range. If the chemical reaction has not taken place the
singlet-state oxygen cannot reach the acceptor bead and the
emission light is not detected. By measuring the intensity of light
it is possible to conclude the efficiency of the chemical
reaction.
[0008] The typical instruments in analytical chemical research
laboratories are the different spectroscopic instruments. Many of
them are utilizing optical region of electromagnetic spectrum. The
two common types of instruments are the spectrophotometers and the
spectrofluorometers. These instruments comprise usually one or two
wavelength dispersion devices, like monochromators. The dispersion
devices make them capable to perform photometric, photoluminescence
and chemiluminescense measurements throughout the optical
spectrum.
[0009] Patent document U.S. Pat. No. 6,538,735 describes a prior
art device for detecting emission from samples. The principle of
the device is illustrated in FIG. 1. In the device 10 the sample is
illuminated by high intensity light produced by a light source 12
such as a laser diode. The light transmitted via a fibre bundle 20
activates the sample, which converts the activation light into
emission light upon biomolecular binding occurrence. The emitted
light is transmitted via a fibre bundle 24 to a detector 41, such
as a photomultiplier tube, which detects and measures the amount of
light after activation ceases. The fibre bundles that transmit
light at the activation and emission wavelength bands are combined
such that the common end of the bundle directly above the well
includes both fibre types. The fibres may be combined e.g.
coaxially. The system can also include a band-pass filter 36 on the
emission side, which eliminates extraneous light, including light
corresponding to the activation wavelength band. The system can be
used in assays based on Amplified Luminescent Proximity Homogeneous
Assay technique. The amount of light produced by the sample is
proportional to the concentration of an analyte in the sample.
[0010] The activation wavelength is between 670 to 690 nm. The
light can be generated by employing a high-intensity laser as the
activation source, emitting in the preferred wavelength region. The
light emitted from the sample has a wavelength band between about
520 nm and 620 nm. This range is at a shorter wavelength than that
of the activation wavelength band. The device may include a shutter
that prevents light from entering the detector while the laser
diode is active, and a filter may prevent light outside the emitted
wavelength band from entering the detector.
[0011] The emitted signal of the AlphaScreen measurement is weak,
and the measurement is sensitive to changes in the environment.
Therefore it is difficult to achieve an efficient and accurate
apparatus for the AlphaScreen measurement. Therefore there are
certain problems related to the prior art arrangements, especially
if several types of measurements are performed with same
equipment.
[0012] The described prior art arrangement of FIG. 1 uses a coaxial
optical cable for transmission and detection. When the
cross-section of the cable is used for separate optical wires for
activation and detection the usable cross section area is very
limited. Therefore both the activation light pulse and the emission
light are much attenuated. The attenuation of the activation and
emission radiation naturally degrades the efficiency and accuracy
of the measurements. The attenuation also causes that the
instrument needs more calibration.
[0013] One solution could be using a dichroic mirror for separating
the optical paths of activation and detection beams as is often
used photoluminescence measurements. A prior art arrangement for
providing photoluminescence measurements is described e.g. in
patent document U.S. Pat. No. 6,071,748. However, there would be
further problems if photoluminescence measurement equipment would
be used also for AlphaScreen measurement. Firstly, if a part of the
confocal optics is same for illumination and detection, it can be
optimised for only one of these purposes. When a sensitive
measurement like AlphaScreen is performed it would be important
that the optics would be optimised for both illumination and
detection. Secondly, different type illumination sources are used
in AlphaScreen and photoluminescence measurements, and therefore it
would be necessary to have optical switches for switching the
optical route between two light sources. However, optical switches
and the related optics attenuate radiation and therefore decrease
the efficiency of the measurements. Good quality optical switches
also tend to increase the manufacturing costs of the
instrument.
[0014] A further significant problem relates to the efficiency of
the prior art solutions. In the AlphaScreen measurement it is
advantageous to use relatively long emission and detection times
for each sample. Therefore it takes a long period of time to
provide measurements for a whole sample well plate that has a large
number of samples. And if several types of measurements are made
for the same samples length of the measurement time increases
further. The long measurement time naturally means that the
throughput of the measurement equipment is not very high. And it
also brings a problem that it may be difficult to keep the
environmental conditions, such as temperature, sufficiently stable
during the measurement of a whole sample assay.
SUMMARY OF THE INVENTION
[0015] An object of the present invention is to provide an optical
instrument for laboratory measurements, wherein the described
disadvantages of the prior art are avoided or reduced. The object
of the invention is therefore to achieve a measurement instrument
with improved versatility, accuracy, reliability and/or efficiency
for performing measurements from samples.
[0016] The object of the invention is achieved by providing an
optical measurement instrumentation wherein a sample is activated
and the emission is detected from the activated sample, and between
the activation and detection phases in measuring the sample, a
shift is made in the relative position between the sample and means
directing the activation radiation to the sample as well as in the
relative position between the sample and the means receiving the
emission radiation from the sample. This can be implemented e.g. by
moving the sample assay plate and/or a measuring head between the
activation and emission phases. The invention allows a simultaneous
activation of a first sample and detecting emission from a second
sample.
[0017] The present invention has several advantages over prior art
solutions. Since the activation beam and the emission beam have
separate optical paths it is possible to optimise the optics for
illumination and detection separately. Thus the accuracy and the
efficiency of the measurement are enhanced. The efficiency of the
measurement is also increased by the fact that the activation and
the detection can be performed simultaneously. It is also possible
to perform different types of measurements simultaneously, such as
photoluminescence measurements and AlphaScreen measurements. This
enhances the overall efficiency further. The accuracy of the
measurement is also enhanced by the fact that a whole sample plate
can be measured in minimal time, and the environmental conditions
can thus be kept stable. As a further advantage, very few optical
components are needed for performing sensitive AlphaScreen
measurements and thus optical attenuation is small and
manufacturing is cost of the instrument is moderate.
[0018] An optical measurement instrument according to the invention
for measuring samples, comprising [0019] an illumination source and
directing means for directing radiation from the illumination
source to a sample for activation of the sample, and [0020]
receiving means for receiving emission radiation caused by said
activation of the sample and a detector for measuring the received
emission radiation from the sample, is characterized in that it
further comprises [0021] first shifting means for changing relative
position between the sample and the directing means between the
phases of activation and receiving emission radiation of the
sample, and [0022] second shifting means for changing relative
position between the sample and the receiving means between the
phases of activation and receiving emission radiation of the
sample, wherein the first shifting means and second shifting means
is same or different means.
[0023] A method according to the invention for optical measurement
of samples, wherein [0024] radiation is directed from an
illumination source into a sample for activation of the sample, and
[0025] emission radiation caused by said activation of the sample
is received and detected for measuring the received emission
radiation from the sample, is characterized in that in the method
[0026] relative position between the sample and radiation directing
means is changed between the phases of activation and receiving
emission radiation of the sample, and [0027] relative position
between the sample and the receiving means is changed between the
phases of activation and receiving emission radiation of the
sample.
[0028] Some preferred embodiments are described in the dependent
claims.
[0029] Said "directing means" means may be e.g. a light guide such
as a fibre optic guide, or it may be the illumination source or
part of it. "Directing means" may be any means with a purpose for
directing activation radiation to a sample. Said "receiving means"
may be e.g. a window of a photomultiplier tube or any receiving
means that receive emission radiation from a sample.
BRIEF DESCRIPTION OF THE DRAWINGS
[0030] The described and other advantages of the invention will
become apparent from the following detailed description and by
referring to the drawings where:
[0031] FIG. 1 is a schematic block diagram of a prior art optical
unit of a measurement instrument,
[0032] FIG. 2 is a schematic illustration of optical paths and main
components of an exemplary optical unit for a measurement
instrument according to the invention,
[0033] FIG. 3 is a schematic block diagram including a side view of
an exemplary measurement instrument according to the invention
where several measurement modes are available,
[0034] FIG. 4 is a front view of an exemplary top measurement head
according to the invention,
[0035] FIG. 5 illustrates a top view of a sample assay plate and
shows above the samples an exemplary positioning of the optical
means for the measurement according to the invention, and
[0036] FIG. 6 is a flow diagram illustrating an exemplary method
for performing an optical measurement according to the
invention.
[0037] FIG. 1 was already explained in the description of the prior
art. In the following, the principle of the invention is first
described referring to FIG. 2. Then, an example of a more detailed
implementation is described referring to FIGS. 3 and 4, which
illustrate main parts of an exemplary analyser equipment according
to the invention. Finally, an exemplary method according to the
invention is described referring to FIGS. 5 and 6.
[0038] FIG. 2 illustrates main components and optical paths of an
exemplary optical analyser instrument according to the invention.
This versatile instrument comprises means for performing both
ordinary type photoluminescence measurements as well as
measurements according to the invention for e.g. AlphaScreen
measurements. Next the parts for ordinary photoluminescence
measurement are first described, and then the parts for e.g. the
Alphascreen measurement.
[0039] The instrument comprises an illumination source 211 for the
activation of a sample in a photoluminescence measurement. The
radiation from the lamp 211 is collimated with lens 215 and
directed through an interference filter 214. Different filters can
be selected for different wavelengths. The activation beam is then
focused with a lens 213 to an end of a fibre optic guide 218, which
guides it to an aperture 246 of an optical module. The fibre optic
guide is preferably a bundle of fibres, such as 200 pieces of
fibres with a diameter of 100 .mu.m.
[0040] The activation beam is guided through an aperture 246 of the
optical module and reflected by a dichroic mirror 241 inside the
optical module 240. The activation beam is further directed into
the sample 281 through an aperture of the optical module and a lens
system 223. A part of the illumination light is reflected by a beam
splitter mirror 243 and guided through an aperture into a reference
detector 219 in order to give reference information on the actual
illumination intensity. A beam splitter mirror can be produced e.g.
by forming reflective coating for the mirror to be e.g. stripes or
dots, which cover only a part of the mirror surface.
[0041] The photoluminescence emission beam from the sample 281 is
directed with the lens system 223 through an aperture into the
optical module 240, where it passes the dichroic mirror 241. The
dichroic mirror is can be designed for a certain label so that it
reflects activation wavelength but transmits emission wavelengths.
The emission beam is then divided inside the optical cube into to
two beams by a second mirror 242. The mirror is preferably a
dichroic mirror, which functions as a filter so that a beam with a
wavelength of the first emission is transmitted through the mirror
and focused through an aperture 244 to the first detector 231a. The
beam with a wavelength of the second emission is reflected and
guided focused through another aperture 245 to the second detector
231b. The second dichroic mirror is therefore designed for each
label/pair of labels so that it transmits first emission
wavelengths but reflects second emission wavelengths.
[0042] The first emission beam received from the aperture of the
optical module is collimated with a lens 233a and directed through
an interference filter 234a in order to prevent light with a
wavelength outside the first emission from passing to the first
detector. The first emission beam is then focused with lens 235a to
the first detector 231a. The second emission beam received from
another aperture of the optical module is reflected with a mirror
238 to a lens 233b where the beam is collimated and directed
through a second interference filter 234b in order to prevent light
with a wavelength outside the second emission from passing to the
second detector. The second emission beam is then focused with lens
235a to the first detector 231a. The signals received from the
detectors are then amplified and processed to achieve a value for
the intensities of the first and second emissions. The instrument
may also comprise a bottom measurement head for measuring radiation
below the sample, via lens 263.
[0043] When the instrument of FIG. 2 is used for e.g. AlphaScreen
measurement the activation light is received from the laser source
212AS, and the beam is directed via an optical guide 218AS to a
sample 283. The fibre optic guide is preferably a bundle of fibres,
such as 1000 pieces of fibres with a diameter of 100 .mu.m. One
purpose of the fibre optic guide is to mix the light of the
illumination source in order to avoid an uneven distribution of
activation beam within the sample volume to be measured. The fibre
optic guide is preferably of such a width that it exactly
illuminates the whole sample. If the fibre optic guide has a
smaller or wider diameter than the sample, a lens system can be
used at the end of the fibre optic guide to adjust the diameter of
the activation light beam to illuminate the whole sample.
[0044] The instrument comprises a detector 291 for AlphaScreen
measurements. In this embodiment the detector is a photo-multiplier
tube. The photo-multiplier may preferably be also used for
chemiluminescence measurements. The photo-multiplier tube is in
this example in a slightly tilted orientation. This may be
necessary in order to perform simultaneously different types of
measurements from samples that are near to each other on a sample
plate.
[0045] The detector receives the radiation from the sample 283 via
an aperture of a disk 290. The radiation reaches the window 293 of
the photo-multiplier tube, and after penetrating through the window
the radiation reaches the active surface of the photo-multiplier
tube. The block 292 includes the preamplifier and other related
electronics for the photo-multiplier tube for measuring the
intensity of the received radiation.
[0046] It is advantageous in the AlphaScreen measurement that the
detector is near to the sample, and the radiation has a clear,
direct path from the sample to the detector. Thus the attenuation
of the emission radiation is negligible. It is also possible to
achieve low attenuation for measuring chemiluminescence emission
beam the described instrumentation. The advantages of the invention
become more apparent in the following more complete example of an
optical instrument according to the invention.
[0047] The present invention is implemented by e.g. first activing
a certain sample and then shifting the position of the sample plate
in relation to the illumination directing means 218AS and
photo-multiplier tube 291, 293 in order to measure the emission.
There is a processor-controlled motor 299 for performing the
shifting of the sample plate between illumination and detection
phases of each sample.
[0048] It would also be possible to include more than one
illumination source and detector for the measurement according to
the invention. Using several illumination source-detector pairs
would naturally increase the measurement efficiency further as
simultaneous activation and detection of several samples would be
possible. It would also be possible to have two or more adjacent
detectors for detecting sequentially emission of same samples. Thus
a sample would be first activated, and after shifting the sample
plate/measurement head the emission would be first detected with a
first detector, and after further shifting the sample
plate/measurement head the emission of the same sample would be
detected with a second detector etc. This way a more accurate
measurement result would be achieved.
[0049] FIG. 3 illustrates in more detail an exemplary optical
instrument according to the invention. Especially, an
implementation of an instrument for several alternative measurement
modes is illustrated in more detail. This shows how the present
invention allows an effective combination of measurement modes in a
single equipment.
[0050] The instrument of FIG. 3 has a top measurement head 320,
which includes components for providing an activation beam and for
detecting emissions from above the sample. The instrument has also
an optional bottom measurement head 360, which includes components
for providing an activation beam and for detecting emissions from
below the sample. The means according to the present invention for
directing activation and/or detection to samples can be included in
the top and/or bottom measurement head. The instrument further
comprises a sample platform 380, which has means for moving a
sample tray 389 in order to position successive samples 381 into
the measurement locations. There may also be means provided for
adjusting the vertical position of the sample platform relative to
the top and bottom measurement heads.
[0051] The instrument comprises a laser source 312AS for e.g.
AlphaScreen measurements The laser source is advantageous due to
its high efficiency on a narrow range of wavelengths. However, also
other illumination sources are applicable, such as Xenon or halogen
lamp used with a filter. The light of the laser source 312AS is
guided in an optical guide 318AS directly to the sample. According
to the present invention the relative position between the sample
tray and the measurement head is changed between the illumination
and detection phases of samples.
[0052] The instrument comprises a detector 391 for detecting the
emission signal in AlphaScreen measurements. The detector may also
be used for chemiluminescence measurements. In this embodiment the
detector is a photo-multiplier tube. The detector receives the
radiation from the sample via an aperture of a disk 390. The
AlphaScreen detector is in front of the photoluminescence
components, and thus the AlphaScreen measurement is made from a
sample which is more on the front, whereas the photoluminescence
measurement is made from a sample which is more on the back in FIG.
3. The fibre optic guide providing the laser activation is located
between the photomultiplier tube 391 and the optics for the
photoluminescence measurement. Thus a photoluminescence measurement
and an AlphaScreen/chemiluminescence measurement can be performed
simultaneously from different samples.
[0053] The detector 391 can be used in analogue mode or digital
mode, or if the properties of the photo-multiplier tube allow, both
modes may be used simultaneously. The preamplifier and other
related electronics for the photo-multiplier tube are located in a
housing 392 above the photo-multiplier tube.
[0054] The aperture discs may be changeable so that different size
apertures can be used with different sample plates. They may
preferably be equipped with machine readable codes, such as bar
codes, so that the processor of the equipment can check with a code
reader, which type of aperture disk is installed. This way it can
be certified that a correct type of aperture disk is used for each
measurement. The bar code reader or related electronics are not
shown in FIG. 3.
[0055] The instrument may also comprise a thermo plate 390 for
keeping the temperature of the samples constant during the
AlphaScreen measurements. The upper measurement head 320 or the
sample platform 380 may be vertically shifted in order to have the
thermo plate tightly between the assay and the upper measurement
head.
[0056] Next the components for performing other types of
measurements are shortly described. The instrument according to
FIG. 3 has another illumination source 312a for providing
activation in photoluminescence measurements. The illumination
source 312a includes a pulse lamp, and the optical energy of each
pulse is preferably equal. The activation beam generated by the
pulse lamp is collimated with a lens 315 and directed through an
interference filter 314. The filter is placed on a filter slide, so
that the activation filter to be used in a measurement can be
selected from several filters. The activation beam is then focused
to an end of a fibre optic guide 318, which mixes the activation
beam and guides it to an aperture of an optical module 340a, which
is located behind the photo-multiplier tube. The optical module 340
and the lens system 323 directs the activation beam into the sample
381.
[0057] The equipment may also include a further pulse lamp 312b,
311b, which may be a low power lamp, e.g. for simultaneous
photometric measurements. The instrument has an optical fibre guide
312a for guiding the light from the second lamp. The light can be
distributed for the photometric measurement into three filters
314h, 314j and 314k with fibre branches 377h, 377j and 377k. After
filtering, the beams are collimated into ends of three optical
fibre cables 378, which are led to the bottom measurement head for
the photometric measurement. The light beams from the optical
cables 378 are focused to three samples 384 with a lens system 379
including lenses for each three beams. After transmitting through
the samples the beams are measured with three detectors 322d, 322e
and 322f, which are e.g. a photo diodes. The three ends of the
fibre optic cables, three lenses, three simultaneously measured
samples and three detectors are in this case located in a row
perpendicular to the plane of the drawing and thus only one of them
can be seen in the drawing.
[0058] It is also possible to use an instrument with same pulse
lamp for photometrics and photoluminescence measurements. For
example, an optical switch 317 may have an output for an optical
fibre 378a, which leads light from the lamp 312a to the
photometrics measurement optics 379. It is then possible to control
the optical switch either to guide the light for providing
activation for an emission measurement or to guide the light the a
photometric measurement.
[0059] An optical fibre 318T is used for guiding the activation
beam from the optical switch 317 to the optical module 340 of the
top measurement head. An optical fibre 318B is used for guiding the
activation beam from the optical switch 317 to the optical module
350 of the bottom measurement head. The instrument may also have a
further lamp so that different lamps can be selected for providing
the activation beam of the top head and the bottom head. In this
case, a more versatile optical switch system is required.
[0060] The emission beam from the sample 381 is directed with the
lens system 323 into the optical module 340 where the emission beam
is divided into to two beams. A dichroic mirror in the optical
module preferably functions as a filter so that a beam with a
wavelength of the first emission is transmitted through the to the
first detector 331a, and a beam with a wavelength of the second
emission is reflected to the second detector 331b. When the
equipment includes two detectors they may be of different types and
there may be alternative detection modes for a photoluminescence
measurement.
[0061] The first emission beam is collimated with a lens 333a and
directed through an interference filter 334j in order to prevent
light with a wavelength outside the first emission from passing to
the first detector. The first emission beam is then focused with
lens 335a to the first detector 331a. The second emission beam is
reflected with a mirror 338 to a lens 333b where the beam is
collimated and directed through a second interference filter 334k
in order to prevent light with a wavelength outside the second
emission from passing to the second detector. The second emission
beam is then focused with lens 335a to the first detector 331a. The
filters 334j and 334k are located on same filter slide or they may
locate on different filter slides. The filter slide(s) is movable
so that the filters used in the measurement can be selected from a
number of filters with different pass-band wavelengths.
[0062] In an instrument also comprising a bottom measurement head
there are optical switches 337a and 337b for selecting the detected
emission beam from the top or bottom measurement head. An optical
fibre 338a is used for guiding the first emission beam from the
optical module 350 of the bottom measurement head 360 to the
optical switch 337a. Another optical fibre 338b is used for guiding
the second emission beam from the optical module 350 of the bottom
measurement head 360 to the optical switch 337b.
[0063] The signals received from the detectors are amplified and
processed to achieve a measurement value for the intensities of the
emissions. Measurement signals and reference signals are amplified
and read after each activation pulse and signal corrections are
calculated. Basic references are determined with standard solvents
after the analyzer has been assembled. Several emission signals
from a same sample may be digitally integrated. Thus the instrument
is also equipped with electronics for amplifying and processing the
signals from the detectors, as well as electronics for driving the
lamp(s). There is also control electronics provided for controlling
the measurements, such as selecting filter(s), selecting the
optical module(s), controlling optical switch(es), controlling the
position of the sample tray 389 according to the invention for
selecting the sample to be measured, and controlling the positions
of the measurement heads 320 and 360 relative to the sample
platform 380. The main electronics is not shown in FIG. 3, as the
required electronics can be designed by a skilled person in the art
using the teachings of the present invention.
[0064] The photo-multiplier tube and its electronics as well as the
light sources are shown reduced in size compared to other
components in FIG. 3. On the other hand, the optical modules are
shown essentially enlarged in FIG. 3 in order to better illustrate
the optical paths in the instruments. FIG. 4 illustrates the sizes
in a more correct relation.
[0065] FIG. 4 illustrates a front view of an exemplary top
measurement head according to the invention. The measurement head
comprises optics 423 for photoluminescence measurements. It also
comprises a photo-multiplier tube 491 with associated amplifier 492
for receiving and detecting emissions in AlphaScreen and
chemiluminoscence measurements. Between the photoluminescence
optics 423 and photo-multiplier tube 491 there is located a fibre
optic guide 418AS for guiding activation light from a laser source
412AS to a sample in e.g. AlphaScreen measurements.
[0066] Next an example of a measurement method according to the
invention is described referring to FIGS. 5 and 6. FIG. 5
illustrates a top view of a sample assay plate 589. It has sample
wells in a 16.times.24 matrix with 16 rows A-Q and 24 lines (N).
Fibre optic guide 518AS providing activation light for AlphaScreen
measurements is above sample well G-4 in the Figure. The detector
591 for detecting Alphascreen and chemiluminescence emission is
above sample well G-2. Due to the dimensions of the detector there
is one sample well G-3 between the activation fibre and the
detector. However, if dimensions allow, the activation fibre and
detector may also be located above adjacent sample wells. FIG. 5
also shows the position of optics 523 for performing
photoluminescence measurements.
[0067] If the AlphaScreen measurement starts at the position
according to FIG. 5, the sample G-4 is first activated. Then the
sample plate is shifted by one step in the right direction, or
alternatively the measurement head is shifted by one step into the
left direction. After this shifting the sample G-5 is activated.
Next the sample plate or measurement head is again shifted by one
step. After this, the sample G-6 is activated and emission from the
sample G-4 is detected. The activation and detection are preferably
simultaneous. This procedure of shifting, activation and detection
is continuing until the last sample in the row (Nmax=24) is
activated and detected. Then samples in other rows A-Q may be
measured. Said "shift by one step" preferably means a shift of a
distance between two adjacent samples. In some cases the shift may
also be a multiple of said distance.
[0068] FIG. 6 illustrates a flow diagram of an exemplary method
according to the invention for performing an AlphaScreen
measurement of a sample assay plate. When measurement of a sample
assay plate starts, 60, the sample plate and measurement head are
positioned for activating a sample in a first row and first sample
N=1 in the row is to be activated, 61, 62. The sample N=1 is then
activated, 63. There is no sample at the position of the detector
(N-2), so detection 64 is not performed at this phase. In phase 65
it is checked whether all samples of the row are measured. If not,
the sample plate or measurement head is shifted by one step ie. "N"
is increased by one. Then sample 2 in the row is activated, 63, and
N is again increased by one. When N has value 3 there is an
activated sample at the position N-2=1 below the detector. Thus it
is possible to perform simultaneously activation of sample N=3, 63,
and detection of emission from sample N-2=1, 64. Thus the procedure
continues by increasing the value of "N" in steps of one, 66, and
performing simultaneous activation of sample N and detection of
emission from sample N-2. When two last samples in the row are
detected, there is no sample at the position of the laser optic
guide, so activation is not performed in phase 63.
[0069] When all samples in a row are measured, it is checked
whether all rows are measured, 67. If not, the measurement
continues at the next sample row, 68. When all sample rows have
been measured, 67, the measurement procedure is ended, 69.
[0070] In the above examples there is one sample well between the
two samples to be simultaneously activated and detected. However,
it would also be possible to have other number of sample wells
between the two samples, starting from zero.
[0071] In this patent specification the structure of the components
in an optical measurement instrument is not described in more
detail as they can be implemented using the description above and
the general knowledge of a person skilled in the art.
[0072] As mentioned above, an optical instrument includes control
means for performing the optical measurement process. The control
of the measuring process in an optical measurement instrument
generally takes place in an arrangement of processing capacity in
the form of microprocessor(s) and memory in the form of memory
circuits. Such arrangements are known as such from the technology
of analyzers and relating equipment. To convert a known optical
instrument into an equipment according to the invention it may be
necessary, in addition to the hardware modifications, to store into
the memory means a set of machine-readable instructions that
instruct the microprocessor(s) to perform the operations described
above. Composing and storing into memory of such instructions
involves known technology which, when combined with the teachings
of this patent application, is within the capabilities of a person
skilled in the art.
[0073] Above, an embodiment of the solution according to the
invention has been described. The principle according to the
invention can naturally be modified within the frame of the scope
defined by the claims, for example, by modification of the details
of the implementation and ranges of use.
[0074] For example, the invention is described as applied to
AlphaScreen measurements. However, even if the invention has
special advantages when applied to such measurements, the invention
can as well be applied in other types of measurements, in which the
length of life time relating to emission is long, such as >0.5
s, compared to the time required for shifting the sample plate. On
the other hand, although the invention is described above as
applied to a versatile instrument for performing several types of
measurements, the invention can also be applied in more simple
instrumentation for e.g. only one type of measurement, such as
AlphaScreen measurement.
[0075] The present invention offers a possibility to perform
activation and detection of separate samples simultaneously and
thus achieve increased effectiveness of the measurements. However,
the present invention is applicable also in measurements with
sequential activation and detection.
[0076] In the above embodiments laser source has been mentioned as
an illumination source in the inventive arrangement. However, also
other illumination sources are applicable, such as xenon or halogen
pulse lamps together with an optical filter. The above embodiments
have included a photo-multiplier tube as a detector. However, many
other types of detectors are also applicable. The detector may be
e.g. a Charge Coupled Device (CCD) detector or camera. It is
possible to achieve a high efficiency/sensitivity with a cooled
CCD.
[0077] Also, although the invention has been described with
reference to the various microtitration plates it is equally
applicable to any form of sample arrangements like vials, discs or
tubes. The samples may be, except liquids, also in other form, such
as gels and filters.
[0078] Although the invention is described with an arrangement
where light source and detector are located on the top measurement
head, there is no reason why their location on the bottom
measurement head should not work. It is also possible to use
illumination from above and detection from below the sample or vice
versa.
[0079] The present invention has advantages in large number of
applications, such as research and clinical applications.
* * * * *