Abstract: The invention provides a highly sensitive measurement of retardance and slow axis orientation, accurately and instantaneously, across a full two-dimensional image. There are no moving parts and there need not be any electro-optic tuning as part of the measurement. It is ideally adapted to real-time imaging and is well-suited to use with biological and medical samples, including visualizing structures in oocytes. The invention splits a light beam into several beams, which are analyzed using elliptical polarizers and the resultant intensity is measured. It can be constructed using a single pixilated detector, or several detectors, to achieve high spatial resolution when this is desired.

Abstract: A variable optical attenuator including: a birefringent element positioned to separate an input optical signal into two spatially separated, orthogonally polarized beams; a LC modulator positioned to receive the orthogonally polarized beams and selectively alter their polarizations; a reflective element positioned to reflect the beams back through the LC modulator and the birefringent element, wherein the birefringent element recombines orthogonally polarized components of the reflected beams to produce an output optical signal; and a controller coupled to the LC modulator to selectively cause the LC modulator to alter the polarizations of the orthogonally polarized beams, wherein during operation the controller is responsive to a request to variably attenuate the intensity of the output optical signal relative to the intensity of the input optical signal to one of multiple non-zero attenuation settings.

Abstract: A microscope system for determining optical properties of a specimen includes a source of polarized light, a detector for detecting the intensity of light incident thereon, an optical path extending from the source to the detector, a condenser for providing light from the source to the specimen, an objective for receiving light from the specimen, a support for mounting the specimen, a sectored variable retarder mounted in the optical path, and a polarized light analyzer mounted in the path between the sectored variable retarder and the detector. The variable retarder has a multiple sectors. Each sector is individually addressable by a control signal that affects the light retardation characteristics of the sector.

Abstract: A variable optical attenuator including: a birefringent element positioned to separate an input optical signal into two spatially separated, orthogonally polarized beams; a LC modulator positioned to receive the orthogonally polarized beams and selectively alter their polarizations; a reflective element positioned to reflect the beams back through the LC modulator and the birefringent element, wherein the birefringent element recombines orthogonally polarized components of the reflected beams to produce an output optical signal; and a controller coupled to the LC modulator to selectively cause the LC modulator to alter the polarizations of the orthogonally polarized beams, wherein during operation the controller is responsive to a request to variably attenuate the intensity of the output optical signal relative to the intensity of the input optical signal to one of multiple non-zero attenuation settings.

Abstract: A variable optical attenuator including: a birefringent element positioned to separate an input optical signal into two spatially separated, orthogonally polarized beams; a LC modulator positioned to receive the orthogonally polarized beams and selectively alter their polarizations; a reflective element positioned to reflect the beams back through the LC modulator and the birefringent element, wherein the birefringent element recombines orthogonally polarized components of the reflected beams to produce an output optical signal; and a controller coupled to the LC modulator to selectively cause the LC modulator to alter the polarizations of the orthogonally polarized beams, wherein during operation the controller is responsive to a request to variably attenuate the intensity of the output optical signal relative to the intensity of the input optical signal to one of multiple non-zero attenuation settings.

Abstract: A microscope system for determining optical properties of a specimen includes a source of polarized light, a detector for detecting the intensity of light incident thereon, an optical path extending from the source to the detector, a condenser for providing light from the source to the specimen, an objective for receiving light from the specimen, a support for mounting the specimen, a sectored variable retarder mounted in the optical path, and a polarized light analyzer mounted in the path between the sectored variable retarder and the detector. The variable retarder has a multiple sectors. Each sector is individually addressable by a control signal that affects the light retardation characteristics of the sector.

Abstract: A polarization imaging system and method are disclosed for measuring the magnitude and orientation of retardance in a sample. There are no moving parts and the invention is readily constructed as an imaging system that obtains polarization values at all point in a scene simultaneously. The system first takes an image that records the apparent slow axis orientation and the apparent retardance. However, the apparent retardance is indeterminate by m&lgr;, the wavelength of observation. By recording such images at two wavelengths, and taking note of both the apparent phase and the angular orientation of polarization interference in each of the two cases, the system is able to determine retardance values of 5&lgr; or more at every point without ambiguity. The actual slow axis orientation is determined as well. The determination of retardance value and axis orientation is independent for each point measured, and does not make use of spatial relationships or distributions within the sample.

Abstract: An imaging system is disclosed comprising an illuminator which produces illumination of any desired pure wavelength or of any selected mixture of pure wavelengths simultaneously, which illuminates a sample without spatio-spectral artifacts using illumination optics designed for that purpose; imaging optics, which form an image of the sample at a detector or viewing port; and a detector. This enables imaging the complete spectral image cube for a sample by taking sequential images while illuminating with a series of pure wavelengths, with greater ease and economy than by means of tunable filters, interferometers and the like. It further enables imaging while the sample is illuminated with a precisely controlled mixture of illuminant wavelengths, so that the image presented to the detector is a linear superposition of the sample properties at many wavelengths.

Abstract: A method in which a multiplicative ratio approach is used to remove the effects of the unwanted background fluorescence when making fluoroescence polarization (FP) measurements rather than the conventional subtractive approach, thus preserving both the precision and accuracy of the FP measurements, is disclosed. The method comprises selecting an appropriate multiplicative ratio, then calculating the selected multiplicative ratio using sample measurements. The calculated multiplicative ratio is multiplied by an appropriate value in a standard FP measurement equation or an appropriate value in an equation derived from a standard FP measurement equation. After this, the corrected FP measurement is calculated. When such multiplicative ratios are applied to the appropriate value or values in an FP measurement equation, the effects of background noise can be reduced without decreasing the precision of the FP measurements.

Abstract: An apparatus for performing fluorescence detection of two or more biochemical probes and/or fluorescence measurement of fluorescence intensity at two or more spectral bands of light emitted from at least one sample spot is disclosed. The apparatus simultaneously directs emitted fluorescent light from multiple probes and/or at multiple spectral bands to different spots on a single pixelated detector.

Abstract: A sample having at least one probe is illuminated with at least one beam of excitation light that is linearly polarized along a first axis, thereby effecting fluorescence emission in at least one spectral band. The intensity of a first component of fluorescence emission that is polarized along the first axis, as well as the intensity of a second component of fluorescence emission that is polarized along an orthogonal second axis, is measured for each of said at least one spectral band. These measurements are represented as a measurement vector M. Since there are various fluorescence sources besides the at least one probe which emit some limited amount of light in the characteristic band of the at least one probe, spectral cross-talk results between the at least one probe and the other fluorescence sources.

Abstract: A sample having a plurality of probe molecules is illuminated with at least one beam of excitation light that is linearly polarized along a first axis, thereby effecting fluorescence emission in a plurality of spectral bands. The intensity of a first component of fluorescence emission that is polarized along the first axis, as well as the intensity of a second component of fluorescence emission that is polarized along an orthogonal second axis, is measured for each of said plurality of spectral bands. These measurements are represented as a measurement vector M. Since each probe emits some limited amount of light in the characteristic band of another probe, this results in cross-talk between probes. The measurement vector is therefore corrected using an instrument response matrix A, which is generated by measuring the flux output of control samples which each have only a single probe species.

Abstract: A variable optical attenuator including: a birefringent element positioned to separate an input optical signal into two spatially separated, orthogonally polarized beams; a LC modulator positioned to receive the orthogonally polarized beams and selectively alter their polarizations; a reflective element positioned to reflect the beams back through the LC modulator and the birefringent element, wherein the birefringent element recombines orthogonally polarized components of the reflected beams to produce an output optical signal; and a controller coupled to the LC modulator to selectively cause the LC modulator to alter the polarizations of the orthogonally polarized beams, wherein during operation the controller is responsive to a request to variably attenuate the intensity of the output optical signal relative to the intensity of the input optical signal to one of multiple non-zero attenuation settings.

Abstract: A method in which a multiplicative ratio approach is used to remove the effects of the unwanted background fluorescence when making fluoroescence polarization (FP) measurements rather than the conventional subtractive approach, thus preserving both the precision and accuracy of the FP measurements, is disclosed. The method comprises selecting an appropriate multiplicative ratio, then calculating the selected multiplicative ratio using sample measurements. The calculated multiplicative ratio is multiplied by an appropriate value in a standard FP measurement equation or an appropriate value in an equation derived from a standard FP measurement equation. After this, the corrected FP measurement is calculated. When such multiplicative ratios are applied to the appropriate value or values in an FP measurement equation, the effects of background noise can be reduced without decreasing the precision of the FP measurements.

Abstract: An instrument is disclosed for fluorescence assays which is capable of reading many independent samples at the same time. This instrument provides enhanced throughput relative to single-sample instruments, and is well-suited to use in general fluorescence, time-resolved fluorescence, multi-band fluorescence, fluorescence resonance energy transfer (FRET), and fluorescence polarization. This invention is beneficial in applications such as high-throughput drug screening, and automated clinical testing. Also disclosed are means and methods for a fluorescence polarization measurement which is highly sensitive, inherently self-calibrated, and unaffected by lamp flicker or photobleaching. This fluorescence polarization invention can be practiced on a variety of fluorescence instruments, including prior-art equipment such as microscopes and multi-well plate readers.

Abstract: An instrument is disclosed for fluorescence assays which is capable of reading many independent samples at the same time. This instrument provides enhanced throughput relative to single-sample instruments, and is well-suited to use in general fluorescence, time-resolved fluorescence, multi-band fluorescence, fluorescence resonance energy transfer (FRET), and fluorescence polarization. This invention is beneficial in applications such as high-throughput drug screening, and automated clinical testing. Also disclosed are means and methods for a fluorescence polarization measurement which is highly sensitive, inherently self-calibrated, and unaffected by lamp flicker or photobleaching. This fluorescence polarization invention can be practiced on a variety of fluorescence instruments, including prior-art equipment such as microscopes and multi-well plate readers.

Abstract: An imaging system is disclosed which provides means for obtaining images with essentially diffraction-limited spatial resolution, and can distinguish between several species of probes within a sample. It may be used with fluorescent, luminescent, up-converting reporter, quantum dot, and other types of probes. Two or more exposures are taken through a filter which expresses different filter states, one of which is preferably a relatively neutral state with high transmission for all wavelengths of interest, and the others of which provide predetermined variation in transmission that are preferably sloping or periodic in wavelength. The probe species is identified by the ratio of response at the various filter settings.

Abstract: An imaging system is disclosed comprising an illuminator which produces illumination of any desired pure wavelength or of any selected mixture of pure wavelengths simultaneously, which illuminates a sample without spatio-spectral artifacts using illumination optics designed for that purpose; imaging optics, which form an image of the sample at a detector or viewing port; and a detector. This enables imaging the complete spectral image cube for a sample by taking sequential images while illuminating with a series of pure wavelengths, with greater ease and economy than by means of tunable filters, interferometers and the like. It further enables imaging while the sample is illuminated with a precisely controlled mixture of illuminant wavelengths, so that the image presented to the detector is a linear superposition of the sample properties at many wavelengths.

Abstract: An imaging system is disclosed comprising an illuminator which produces illumination of any desired pure wavelength or of any selected mixture of pure wavelengths simultaneously, which illuminates a sample without spatio-spectral artifacts using illumination optics designed for that purpose; imaging optics, which form an image of the sample at a detector or viewing port; and a detector. This enables imaging the complete spectral image cube for a sample by taking sequential images while illuminating with a series of pure wavelengths, with greater ease and economy than by means of tunable filters, interferometers and the like. It further enables imaging while the sample is illuminated with a precisely controlled mixture of illuminant wavelengths, so that the image presented to the detector is a linear superposition of the sample properties at many wavelengths.

Abstract: An apparatus for performing fluorescence detection of two or more biochemical probes and/or fluorescence measurement of fluorescence intensity at two or more spectral bands of light emitted from at least one sample spot is disclosed. The apparatus simultaneously directs emitted fluorescent light from multiple probes and/or at multiple spectral bands to different spots on a single pixelated detector.