Optics: measuring and testing – By dispersed light spectroscopy – With sample excitation
Reexamination Certificate
2000-01-14
2003-07-22
Smith, Zandra V. (Department: 2877)
Optics: measuring and testing
By dispersed light spectroscopy
With sample excitation
C356S311000, C356S246000
Reexamination Certificate
active
06597450
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to an apparatus and method for optically reading a plurality of samples in a biological or chemical assay, such as a nucleic acid assay. More particularly, the present invention relates to an apparatus and method which controls two or more light sensor bars each including a plurality of light sources to excite a plurality of samples with a different wavelength of light for each of said two or more light sensor bars in a time-staggered fashion to cause each of the samples to fluoresce at different times and a plurality of light detecting ports interconnected to a single detector for each of said two or more light sensor bars to detect the light emitted from each of the samples.
BACKGROUND OF THE INVENTION
In the clinical diagnosis of bacterial diseases, such as tuberculosis, a sample of sputum or other body fluid obtained from the patient is cultured to test for the presence of the particular bacterium of interest. Unfortunately, this is a relatively time-consuming process, generally requiring several days to produce a definitive result. During this time, a patient suspected of having tuberculosis, for example, must be isolated to prevent further spread of the disease.
The advent of nucleic acid probes, which can identify a specific bacterium by testing for the presence of a unique bacterial nucleic acid sequence in the sample obtained from the patient, has greatly increased the speed and reliability of clinical diagnostic testing. A test for the tuberculosis mycobacterium, for example, can be completed within a matter of hours using nucleic acid probe technology. This allows treatment to begin more quickly and avoids the need for long patient isolation times.
In the use of nucleic acid probes for clinical diagnostic purposes, a nucleic acid amplification reaction is usually carried out in order to multiply the target nucleic acid into many copies or amplicons. Examples of nucleic acid amplification reactions include strand displacement amplification (SDA) and polymerase chain reaction (PCR). Detection of the nucleic acid amplicons can be carried out in several ways, all involving hybridization (binding) between the target nucleic acid and specific probes.
Many common nucleic acid probe detection methods involve the use of fluorescein dyes. One known detection method is fluorescence energy transfer. In this method, a detector probe is labeled both with a fluorescein dye that emits light when excited by an outside source, and with a quencher which suppresses the emission of light from the fluorescein dye in its native state. When nucleic acid amplicons are present, the fluorescein-labeled probe binds to the amplicons, is extended, and allows fluorescence emission to occur. The increase of fluorescence is taken as an indication that the disease-causing bacterium is present in the patient sample.
Several types of optical readers or scanners exist which are capable of exciting fluid samples with light, and then detecting any light that is generated by the fluid samples in response to the excitation. For example, an X-Y plate scanning apparatus, such as the CytoFluor Series 4000 made by PerSeptive Biosystems, is capable of scanning a plurality of fluid samples stored in an array or plate of microwells. The apparatus includes a scanning head for emitting light toward a particular sample, and for detecting light generated from that sample. The apparatus includes first and second optical cables each having first and second ends. The first ends of the optical cables are integrated to form a single Y-shaped “bifurcated” cable. The scanning head includes this end of the bifurcated optical cable. The second end of the first optical cable of the bifurcated cable is configured to receive light from a light emitting device, such as a lamp, and the second end of the second cable of the bifurcated cable is configured to transmit light to a detector, such as a photomultiplier tube.
During operation, the optical head is positioned so that the integrated end of the bifurcated optical fiber is at a suitable position with respect to one of the microwells. The light emitting device is activated to transmit light through the first optical cable of the bifurcated optical cable such that the light is emitted out of the integrated end of the bifurcated optical cable toward the sample well. If fluid sample fluoresces in response to the emitted light, the light produced by the fluorescence is received by the integrated end of the optical fiber and is transmitted through the second optical fiber to the optical detector. The detected light is converted by the optical detector into an electrical signal, the magnitude of which is indicative of the intensity of the detected light. This electrical signal is processed by a computer to determine whether the target nucleic acid is present or absent in the fluid sample based on the magnitude of the electrical signal.
In this type of X-Y plate reader apparatus, the reader head must be repositioned for each well. Accordingly, if the microwell array is a standard microwell array having 12 columns of 8 microwells (96 microwells total), the reader head must move 96 times for the entire microwell array to be read. This excessive movement increases the amount of wear and tear experienced by the apparatus. Furthermore, the control system for controlling the positioning of the head reader must be sophisticated enough to ensure that the integrated end of the optical fiber in the reader head is positioned correctly for each microwell so that the readings are taken at identical locations (e.g., the center) of each microwell. If the integrated end of the optical fiber is not aligned correctly with the microwell, the fluid in the microwell may not receive an adequate amount of excitation light and may therefore not fluoresce properly. Furthermore, any fluorescence that does occur may not be completely detected, because that light may not transmit properly into the integrated end of the bifurcated optical fiber. Accordingly, unless the positioning of the head reader is maintained precise for each individual well, erroneous readings may occur.
Another existing type of apparatus is described in U.S. Pat. No. 5,473,437, to Blumenfeld et al. This apparatus includes a tray having openings for receiving bottles of fluid samples. The tray includes a plurality of optical fibers which each have an end that terminates at a respective opening in the tray. The tray is connected to a wheel, and rotates in conjunction with the rotation of the wheel. The other ends of the optical fibers are disposed circumferentially in succession about the wheel, and a light emitting device is configured to emit light toward the wheel so that as the wheel rotates, the ends of the optical fibers sequentially receive the light being emitted by the light emitting device. That is, when the wheel rotates to a first position, a fiber extending from the wheel to one of the openings becomes aligned with the optical axis of the light emitting device and thus, the emitted light will enter that fiber and be transmitted to the opening. The apparatus further include a light detector having an optical axis aligned with the optical axis of the emitted light. Accordingly, if the sample in the bottle housed in the opening fluoresces due to the excitation light, the light emitted from the sample will transmit through the optical fiber and be detected by the detector. The wheel then continues to rotate to positions where the ends of the other optical fibers become aligned with the optical axis of the light emitter and light detector, and the light emission and detection process is repeated to sample the fluid samples in the bottles housed in the openings associated with those fibers.
As with the CytoFluor X-Y plate reader apparatus described previously, the apparatus described in U.S. Pat. No. 5,473,437 uses a single light emitter and a single light detector to test a plurality of fluid samples. However, instead of using a single bifurcated cable as in the X-Y plate reader app
Andrews Jeffrey P.
Failing Frank L.
Hansen Timothy
O'Keefe Christian V.
Pope Willard C.
Becton Dickinson and Company
Kiang Allan M.
Smith Zandra V.
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