The Perkin-elmer Corporation

Perseptive Biosystems Inc.

Applera Corporation

Mds Inc.

Ensemble Therapeutics Corporation

Lawrence A Haff

Timothy M Woudenberg

Will Bloch

Enrico G Picozza

John Girdner Atwood

David H Tracy

Robert P Ragusa

Paul G Saviano

Igor P Smirnov

Joseph DiCesare

James M Coull

Michael Egholm

Caifu Chen

Jennifer M Campbell

Richard W Noreiks

Andrew J Tomlinson

Joshua A Bittker

Larry W McLaughlin

Cheryl E Murphy

The invention provides turnover probes for use in a variety of detection assays, for example, nucleic acid detection assays. In addition, the invention provides assays, for example, nucleic acid detection assays, using such turnover probes.

Turn over probes and use thereof for nucleic acid detection

A sample plate for a MALDI process is provided which comprises an electrically conductive substrate having a hydrophobic coating whose thickness and hydrophobic character can be modified by changing the coating substrate and/or the concentration of the substrate. Different coating substances that have provided optimal performance of the sample plate for reproducible deposition and analysis by MALDI-MS and MALDI-MS/MS processes of analyte mixtures include synthetic waxes such as paraffin compositions, lipids, organic acids, silicon-containing compounds, silica polymers and natural waxes. Metal polishes that have been used to clean and regenerate plate surfaces have also provided a sample plate that has optimal performance for reproducible deposition and analysis by MALDI-MS and MALDI-MS/MS processes of analyte mixtures.

MALDI plate and process for making a MALDI plate

An asynchronous thermal cycling protocol for nucleic acid amplification uses two primers with thermal melting temperatures different by about 10 to 30&#xb0; C. After the higher melting primer has annealed and polymerase mediated extension, the uncopied, single-stranded target sequence may be hybridized and detected by a probe. DNA probes may be cleaved by the exonuclease activity of a polymerase. The probe may be a non-cleaving analog such as PNA. When a probe is labelled with a reporter dye and a quencher selected to undergo energy transfer, e.g. FRET, fluorescence from the reporter dye may be effectively quenched when the probe is unbound. Upon hybridization of the probe to complementary target or upon cleavage while bound to target, the reporter dye is no longer quenched, resulting in a detectable amount of fluorescence. The second, lower-melting primer may be annealed and extended to generate a double-stranded nucleic acid. Amplification may be monitored in real time, including each cycle, or at the end point. The asynchronous PCR thermal cycling protocol can generate a preponderance of the PCR amplicon in single-stranded form by repetition at the end of the protocol of annealing and extension of the higher melting primer.

Asynchronous primed PCR

Apparatus and method for performing a nucleic acid amplification reaction and preferably a polymerase chain reaction (PCR) in a reaction mixture in at least one capillary tube. Several different embodiments are disclosed. One embodiment cycles a sample through a capillary tube loop passing through two thermostatted fluid baths. Another embodiment has the capillary tube routed alternatingly between two heat exchangers to that the sample makes only one pass through the tube. Other embodiments maintain the heat exchangers stationary and translate the samples between them. Still further embodiments maintain the samples stationary and either automatically translate or rotate the heat exchangers past the samples contained within the capillary tubes to perform the thermal cycles necessary for the amplification reaction.

Nucleic acid amplification reaction apparatus and method

The invention relates to methods for determining sequence information in polynucleotides by combining the recent disparate technologies of mass spectrometry and polynucleotide hybridization, amplification, extension and/or ligation techniques. Broadly, in a first step, the method for determining sequence information in a sample polynucleotide includes hybridizing with a sample nucleotide one or a mixture of oligonucleotide probes having a nucleotide sequence complementary to a portion of the sample polynucleotide, thereby forming a complex. Then, in a second step, the complex is contacted with at least a member selected from the group consisting of nucleosides, dideoxynucleosides, polymerases, nucleases, transcriptases, ligases and restriction enzymes to alter at least a subset of said oligonucleotide probes. In a third step, the method provides for determining the molecular weight of at least the subset of altered probes by mass spectrometry and infering the sequence information of the sample polynucleotide therefrom.

Methods for determining sequences information in polynucleotides using

Nucleic acid amplification reaction apparatus

A complete in situ PCR system for amplification of nucleic acids contained in a prepared cell or tissue sample. The containment system for the PCR sample comprises a glass microscope slide, a specimen sample containing the target nucleic acid sequence mounted on the slide, a flexible plastic cover over the sample, and a retaining assembly fastened to the slide and to the cover to retain and seal a reaction mixture against the sample during thermal cycling. The retaining assembly includes a rigid ring on a rim portion of the cover, a cross beam having spaced parallel rails joined by opposite flat ends, and a pair of clips which are pressed over the ends and opposite sides of the slide to fasten the cross beam and cover to the slide.

Assembly system for in situ per amplification

In situ PCR amplification system

A kit and method for reduction of primer-independent background signals in a DNA replication/detection assay, particularly in a direct in situ PCR amplification/detection assay, for a target DNA sequence in DNA sample, comprising pretreating the DNA sample, containing the target DNA sequence to be replicated/amplified and assayed, with DNA polymerase and one or more dideoxynucleoside triphosphates prior to conducting the replication/amplification steps.

Method for reducing background signals in DNA replication/detection

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Wilton, CT, United States of America

Lawrence A Haff