Abstract: A method for making a model for the folded structure of a set of proteins from an evolutionary analysis of a set of aligned homologous protein sequences was claimed in Ser. No. 07/857,224. The instant application concerns methods for using these models. The first method is used to confirm or deny a hypothesis that two proteins are homologous, and is comprised of comparing a predicted structure model for one family of proteins with a predicted structure model for a second family of proteins, or an experimental structure for the second family, and deducing the presence or absence of homology based on the presence or absence of structural similarity flanking key residue motifs in the polypeptide sequence.

Abstract: The invention concerns the use of peptides of general formula R1-L-Tyr-L-Arg-R2 in which R1=H or a R3-C=0 with R3=a C1 to C20 alkyl chain, linear or branched, saturated or unsaturate, hydroxylated or not, or with R3=an aryl, aryl-alkyl or alkyloxy or aryloxy or arylalkyloxy group, and in which R2=OH or a O-R4 group with R4=a C1 to C20 alkyl chain, or R2=a NH2, NHX or NXX group with X=a C1 to C4 alkyl chain. The peptides have a soothing effect on the skin, including by topical application, and attenuates the effects of benign skin sores (after shave irritation, sunstroke, frostbite, chaps, depilation). They are used in acceptable cosmetic excipients and in effective in vivo concentrations.

Abstract: The present invention discloses novel methods and apparatuses for mass spectrometry. In the methods and apparatuses of the invention, ions are accumulated in an ion reservoir and dissociated with coherent radiation prior to mass analysis. These methods and apparatuses are amenable to mass spectrometric analysis of biomolecules and are particularly usefuil for the sequencing of oligonucleotides, peptides and oligosaccharides.

Abstract: The present invention relates to a method of identifying a molecule of a molecule-substrate complex, wherein the molecule is covalently attached directly to a substrate or indirectly by means of a linking moiety, comprising: (a) bombarding the molecule-substrate complex with energized particles to cleave the molecule from the molecule-substrate complex; and (b) determining the molecular weight of the cleaved molecule by means of mass spectrometry. The inventive method may further comprise irradiating the cleaved molecule with photons.

Abstract: A method for accomplishing a plurality of combinatorial processes in parallel using a microelectronic and fluidic array (device array) having micron-sized reservoirs, connecting microchannels and reaction cells etched into a substrate. The device array has a top feedthru plate, a center distribution plate and a bottom cell plate. The top feedthru plate serves as a cover for the device array and contains apertures selectively positioned above the reservoirs located in the center distribution plate. The center distribution plate includes a plurality of micron sized reservoirs, microchannels, reservoir feeds, cell feeds and overflow feeds for the distribution of reagent fluids to the reaction cells located in the bottom cell plate. The detachable bottom cell plate serves as a microlaboratory tray of reaction cells. Once the proper reagents or other materials are introduced into the reaction cells, the bottom cell plate is decoupled from the device array and removed for incubation or analysis.

Abstract: The binding sites of binding proteins and their binding partners are characterized, at the individual amino acid level, by a combination of tritium exchange labeling and sequential degradation and analysis of tritiated fragments under slowed exchange conditions.

Abstract: The invention relates to the staining of amine-containing polymers, including including peptides, polypeptides, and proteins, in gels and on solid supports, using complexes of europium (3+).

Abstract: A sample presentation device, with a surface-bound complex including at least one molecule which chemically modifies a biomolecule, is prepared and exposed to a biomolecule. The molecular weights of the chemically modified biomolecule is then determined by mass spectrometry.

Abstract: The present invention provides methods whereby the positions of peptide amide groups that are labeled with a heavy hydrogen in a polypeptide or protein can be localized at high resolution. The methods are useful for determining which peptide amide groups in a polypeptide or protein are accessible to solvent, mapping the binding site and/or binding surface of a binding protein, and/or studying allosteric or other conformational changes in a polypeptide or protein which alter the rates at which certain peptide amide hydrogens exchange with solvent.

Abstract: A disposable high density optically readable polydeoxynucleotide array with integral fluorescence excitation and fluorescence emission channels is described. The compact array size allows integration into several types of interventional devices such as catheters, guidewires, needles, trocars and may be used intraoperatively. Highly sensitive monitoring of the metabolic and disease pathways of cells in vivo under varying chemical, genetic and environmental conditions is afforded.

Abstract: The present invention provides a method for producing a peptide thiol ester using fluoren-9-ylmethoxycarbonylamino acid (Fmoc-amino acid). The method is for peptide synthesis and involves (1) using and removing the Fmoc group bound as the protective group to the amino group of amino acid, fixed on a resin via the thiol ester bond, a specific reagent is used to remove an Fmoc group from the amino acid thiol ester resin; (2) adding Fmoc-amino acid to the Fmoc-freed resin and then removing the Fmoc group, repeatedly, to prepare the Fmoc-peptide thiol ester resin; and (3) treating sequentially, the Fmoc-peptide thiol ester resin with a cleavage reagent and with a reagent capable of removing the Fmoc group.

Abstract: A modified Edman degradation process is used to obtain compositional tags for proteins. The Edman degradation chemistry is separated from amino acid analysis, circumventing the serial requirement of the conventional Edman process. Multiple cycles of coupling and cleavage are performed prior to extraction and compositional analysis of amino acids. The amino acid composition information is used to search a database of known protein or DNA sequences to identify the sample protein.

Abstract: Method is described for sequencing polypeptides by forming peptide ladders comprising a series of polypeptides in which adjacent members of the series vary by one amino acid residue and determining the identity and position of each amino acid in the polypeptide by mass spectroscopy.

Abstract: An improved biologic electrode array and methods for manufacturing and using the same. In one aspect, a matrix of electrodes each coupled to a respective sample-and-hold circuit is provided. The electrodes and sample-and-hold circuits are integral and form an array within a single semiconductor chip, such that each sample-and-hold circuit may be loaded with a predefined voltage provided by a single, time-shared digital-to-analog converter (DAC). Further, all of the sample-and-hold circuits may be accessed through a multiplexer which may be scanned through some or all of the electrode locations. Each sample-and-hold circuit may comprise a capacitor and one or more transistor switches, the switch(es), when closed, providing electrical communication between the capacitor and a source line formed in the matrix.

Abstract: A method of screening an agent for potential use in the treatment of Alzheimer's disease, comprises reacting, in the presence of the agent, tau protein with a suitable sulphated carbohydrate under appropriate conditions to form filaments, and monitoring for the presence of filaments. Tau protein and sulphated carbohydrate, e.g. sulphated glycosaminoglycan, will react under appropriate conditions to form filaments, either paired helical filaments or straight filaments. If filament formation is affected when the reaction is carried out in the presence of an agent being screened, this is possibly due to an interfering, inhibiting or blocking effect of the agent. An agent which inhibits assembly of PHFs in vitro may also have an inhibiting effect in vivo and thus have potential therapeutic value in delaying the dementing effects of Alzheimer's disease. The invention can thus provide a screen to identify agents worthy of further investigation for use in the treatment of Alzheimer's disease.

Abstract: The present computer-implemented process involves a methodology for determining properties of ligands which in turn can be used for designing ligands for binding with protein or other molecular targets, for example, HIV targets. The methodology defines the electrostatic complement for a given target site and geometry. The electrostatic complement may be used with steric complement for the target site to discover ligands through explicit construction and through the design or bias of combinatorial libraries. The definition of an electrostatic complement, i.e., the optimal tradeoff between unfavorable desolvation energy and favorable interactions in the complex, has been discovered to be useful in ligand design. This methodology essentially inverts the design problem by defining the properties of the optimal ligand based on physical principles.

Abstract: Methods for performing rapid and accurate thermocycling on a sample are disclosed. Use of non-contact heating and cooling sources allows precise temperature control with sharp transitions from one temperature to another to be achieved. A wide range of temperatures can be accomplished according to these methods. In addition, thermocycling can be performed without substantial temperature gradients occurring in the sample. Apparatus for achieving these methods are also disclosed. A method for pumping a sample through microchannels on a microchip using a non-contact heat source is also disclosed.

Abstract: A method for detecting cyclization of acyclic compounds is disclosed. In particular, the invention relates to a method of screening for macrocyclic peptidase inhibitors, and is useful for screening a combinatorial library of compounds.

Abstract: A non-invasive method for the determination of oxidative stress in a patient by urinalysis is disclosed. The method comprises quantifying the level of o,o'-dityrosine in a sample of the urine of said patient and comparing with the corresponding level of said compound in a normal or control sample, whereby a substantially elevated level of said o,o'-dityrosine is indicative of oxidative stress in said patient.

Abstract: The invention includes compositions comprising and methods of using 1,2-indanedione derivatives for detecting an amine compound such as an amino acid. Methods of detecting and recording the pattern of a fingerprint on a surface are also included, as are a kit for detecting an amine compound such as a constituent of a fingerprint. The invention further includes a device for developing a fingerprint and a method of making 1,2-indanedione derivatives.

Abstract: The invention is directed to a method and device for simultaneously testing a sample for the presence, absence, and/or amounts of one or more a plurality of selected analytes. The invention includes, in one aspect, a device for detecting or quantitating a plurality of different analytes in a liquid sample. The device includes a substrate which defines a sample-distribution network having (i) a sample inlet, (ii) one or more detection chambers, and (iii) channel means providing a dead-end fluid connection between each of the chambers and the inlet. Each chamber may include an analyte-specific reagent effective to react with a selected analyte that may be present in the sample, and detection means for detecting the signal. Also disclosed are methods utilizing the device.

Abstract: A noninvasive method for the determination of oxidative stress in a patient is disclosed. The method comprises quantifying the levels or relative distribution of a pair of compounds, o,o'-dityrosine and o-tyrosine, in a sample of the patient's urine and comparing with the corresponding levels or relative distribution of the compounds in a normal or control sample.

Abstract: The invention provides a method for synthesizing various chemicals onto solid supports, cleaving the synthesized compounds and preparing samples for analysis. In one exemplary embodiment, the invention provides a device comprising a housing which defines an enclosure. A plate having a plurality of wells is received into the enclosure. Each of the wells has a bottom end and at least some of the wells have a hole in the bottom end. A pressure source is in fluid communication with the holes in the bottom ends of the wells. In this manner, a fluid may be maintained within the wells by application of pressure from the pressure source.

Abstract: A system and method for detection, identification, and monitoring of submicron sized particles, the method including the steps of collecting a sample, extracting existing submicron particles from the collected sample based on density, purifying the extracted submicron particles by concentrating the extracted submicron particles based on size, and, detecting and identifying the purified extracted submicron particles based on size and density thereby determining submicron particles present in the collected sample. The submicron particles detected and identified include viruses and virus-like agents such as prions. Thus, virus and virus-like agents can be detected and identified based only on their physical properties without the use of biochemical reagents or assays. A system for carrying out the method of detection and identification of these particles is also disclosed.

Abstract: A process is disclosed for sequencing proteins or peptides from the C-terminal end. The process comprises the steps of reacting the peptide or protein with an alkyl acid anhydride to convert the carboxy-terminal thereof into oxazolone, liberating the C-terminal amino acid by reaction with acid and alcohol or with ester, and identifying the liberated amino acid or amino acid derivative.

Abstract: The present invention provides methods of electrophoretically separating macromolecular species, as well as compositions and systems useful in carrying out such methods. Specifically, the methods of the present invention comprise providing a substrate that has at least a first capillary channel disposed therein. The surface of the channel has a first surface charge associated therewith, and is filled with a water soluble surface adsorbing polymer solution that bears a net charge that is the same as the charge on the capillary surface.

Abstract: An array is created by moving a dispenser toward a solid support until a tip of the dispenser touches the support, withdrawing the tip from the surface and releasing a drop of 5 nanoliters or less in an area smaller than 1 mm.sup.2 to create an array of at least 100 spots.

Abstract: This invention pertains to methods of determining the presence and/or quantity of homocysteine in a sample containing other thiol-containing compounds. The methods of this invention involve modifying the homocysteine to facilitate the separation of homocysteine from cysteine. An assay protocol comprises adjusting the conditions of the sample suspected of containing homocysteine so that homocysteine forms homocysteine thiolactone, separating the homocysteine thiolactone from free thiol-containing compounds present in the sample, including cysteine, reconverting the homocysteine thiolactone to homocysteine, and determining the presence and/or quantity of homocysteine in the sample by conventional means.

Abstract: A method for correlating a peptide fragment mass spectrum with amino acid sequences derived from a database is provided. A peptide is analyzed by a tandem mass spectrometer to yield a peptide fragment mass spectrum. A protein sequence database or a nucleotide sequence database is used to predict one or more fragment spectra for comparison with the experimentally-derived fragment spectrum. In one embodiment, sub-sequences of the sequences found on the database which define a peptide having a mass substantially equal to the mass of the peptide analyzed by the tandem mass spectrometer are identified as candidate sequences. For each candidate sequence, a plurality of fragments of the sequence are identified and the masses and m/z ratios of the fragments are predicted and used to form a predicted mass spectrum.

Abstract: A carboxy terminal protein sequencing process is disclosed which utilizes diphenyl phosphoroisothiocyanatidate and a heterocyclic amine to produce a thiohydantoin derivative of the C-terminal amino acid. The derivative is readily cleaved. The method is useful to sequence through all of the 20 naturally occuring amino acids.

Abstract: A method for C-Terminal degradation of peptides and proteins involves forming a thiohydantoin derivative of the C-Terminal amino acid and then cleaving the derivatized amino acid by reaction with methoxide or methiolate ions.

Abstract: The present invention generally provides microfluidic devices which incorporate improved channel and reservoir geometries, as well as methods of using these devices in the analysis, preparation, or other manipulation of fluid borne materials, to achieve higher throughputs of such materials through these devices, with lower cost, material and/or space requirements.

Abstract: The present invention provides for an efficient and novel method for the C-terminal sequencing of proteins or peptides by use of acetyl chloride or phosphoryl chloride by reaction with a suitable isothiocyanate for derivitation of the carboxy terminus to a thiohydantoin amino acid derivative, under acidic conditions. Cleavage of the derivatized thiohydantoin amino acid may occur by use of thiocyanic acid and acetic acid in water and also by the novel means using a buffer and a potassium or sodium thiocyanate or potassium or sodium dithionite reagent. The present invention also provides for an novel and efficient means for the C-terminal sequencing of proteins or peptides by a two or three step process which comprises first reacting the peptide or protein with an acid chloride reagent, such as acetyl chloride, or phosphoryl chloride.

Abstract: An improved biologic electrode array and methods for manufacturing and using the same. In one aspect, a matrix of electrodes each coupled to a respective sample-and-hold circuit is provided. The electrodes and sample-and-hold circuits are integral and form an array within a single semiconductor chip, such that each sample-and-hold circuit may be loaded with a predefined voltage provided by a single, timeshared digital-to-analog converter (DAC). Further, all of the sample-and-hold circuits may be accessed through a multiplexer which may be scanned through some or all of the electrode locations. Each sample-and-hold circuit may comprise a capacitor and one or more transistor switches, the switch(es), when closed, providing electrical communication between the capacitor and a source line formed in the matrix.

Abstract: A method is presented for predicting the folded structure of proteins that comprises obtaining an alignment of the sequences of a set of homologous proteins, using patterns of conservation and variation of the sequence between proteins with clearly defined evolutionary relationships to assign positions in the alignment to the surface of the folded structure, the inside of the folded structure, active site, or parsing segments, assigning secondary structures by identifying periodicity in said assignments, and then assembling the secondary structural units into a globular form using distance constraints imposed by disulfide bridges, active site assignments, and covariation analysis.

Abstract: A method, apparatus and article for sequencing polypeptides and proteins ("macromolecules") utilizes a sequencing packet comprised of a plurality of substrates having two different derivatization chemistries within a forarninous container. The sequencing packet is disposed within a reaction chamber in a cartridge assembly having an inlet and outlet. An unknown target macromolecule, and preferably a control macromolecule are bonded to the substrates. The macromolecule and control are then simultaneously sequenced. The control serves to monitor the efficacy of the sequencing process.

Abstract: The present invention provides a novel internal standard for amino acid sequencing which contains a peptide consisting of unnatural amino acid residues, such as ornithine, norvaline, norleucine and .alpha.-aminobutyric acid, that is capable of being sequenced simultaneously with an unknown peptide or protein without interfering with the analysis. The internal standard peptide has an amino acid sequence containing at least two different unnatural amino acid residues having retention times distinct from the corresponding retention times for natural amino acid residues. Information derived from the sequencing of the internal standard allows determination of repetitive yield, lag, N-terminal blockage and discrimination between blank cycles caused by missed injection and blank cycles caused by faulty delivery of chemicals during the sequencer reactions.

Abstract: Disclosed are methods that can be used to (1) measure the level of polysaccharide in a sample; (2) measure the ability of a compound to degrade a polysaccharide; (3) measure the ability of a compound to modulate polysaccharide synthesis; and (4) identify or distinguish a polysaccharide, and hence organism, for diagnostic purposes in clinical medicine or research. The invention stems from Applicant's discovery that polysaccharides have multiple binding sites for polysaccharide binding moieties (PBM, e.g., wheat germ agglutinin (WGA)). In each method, one PBM links the polysaccharide to a substrate, and a tagged PBM is used to detect the polysaccharide. All of these methods can be carried out rapidly and quickly in the wells of a microtiter plate, thus permitting high through-put screening of samples or test compounds.

Abstract: System for analyzing a protein sample, comprising a reactor vessel, a chromatographic column, a mass spectrometer, and a computer system. The reactor vessel comprises an enzyme activity capable of digesting the protein sample in order to provide a plurality of peptide digests, an inlet port for receiving the protein to be digested, and an exit port for discharging the peptide digests. The chromatographic column comprises a chromatographic medium capable of chromatographically fractionating the peptide digests as the peptide digests are eluted through the column, wherein the chromatographic column comprises an inlet port for receiving the peptide digests, said inlet port being in flow communication with the exit port of the reactor vessel, and wherein the chromatographic column comprises an exit port for discharging an effluent comprising the chromatographically fractionated peptide digests.

Abstract: A microscale fluid handling system that permits the efficient transfer of nanoliter to picoliter quantities of a fluid sample from the spatially concentrated environment of a microfabricated chip to "off-chip" analytical or collection devices for further off-chip sample manipulation and analysis is disclosed. The fluid handling system is fabricated in the form of one or more channels, in any suitable format, provided in a microchip body or substrate of silica, polymer or other suitable non-conductive material, or of stainless steel, noble metal, silicon or other suitable conductive or semi-conductive material. The microchip fluid handling system includes one or more exit ports integral with the end of one or more of the channels for consecutive or simultaneous off-chip analysis or collection of the sample. The exit port or ports may be configured, for example, as an electrospray interface for transfer of a fluid sample to a mass spectrometer.

Abstract: A method and apparatus for sequencing polypeptides, including a continuous flow reactor which may include a sample bearing membrane strip.

Abstract: A first reactant is immobilized i.e. in a porous matrix (50), adjacent a sample electrode (46) within a reaction chamber. Energizing of the electrode (46) electrophoretically attracts a mobile second reactant and/or electrolytically induces appropriate reaction conditions to enhance reaction of the first and second reactants. Polarity reversals between the sample electrode (46) and remote electrodes (38), (42), (44) cause unreacted second reactant and/or by-products to migrate away from the immobilized first reactant. The techniques are useful for sequential chemical reactions such as sequencing or construction of proteins, polysaccharides and nucleic acids where cyclical additions and removals of reactants are required. The techniques are amenable to automated micro and nano scale construction and operation and allow direct electrophoretic (38) interfacing with chromatographic, HPCE and mass spectrophotometric equipment.

Abstract: Isolated DNA molecules comprising a nucleotide sequence encoding murine son of sevenless gene 1 (mSOS1) polypeptide and comprising a nucleotide sequence encoding murine son of sevenless gene 2 (mSOS2) polypeptide are disclosed, as well as isolated mSOS1 polypeptide and isolated mSOS2 polypeptide, and diagnostic methods using the same.

Abstract: A method of detecting nucleic acids, proteins, or protein nucleic acid complexes. The method includes binding an enzyme, such as phosphatase, to a specimen of the nucleic acid, protein, or protein nucleic acid complex. The enzyme is then reacted with a fluorescein derivative phosphate ester to obtain a fluorescein derivative phosphate ester hydrolysate. The hydrolysate is then irradiated with excitation light, and the emitted fluorescein is detected.

Abstract: A method for chemically generating a set of N-terminally truncated peptides (ladder), suitable for analysis by mass spectrometry (MS), from a peptide or protein using carbon disulfide (CS.sub.2). The method consists of: 1) a coupling step employing CS.sub.2, a tert-amine catalyst, and a solvent under conditions that maximize the yield of dithiocarbamyl-peptide, and 2) a cleavage step employing an acid, which catalyzes two alternate pathways, removal of the N-terminal residue and regeneration of the starting peptide, with no other modification. The simultaneous operation of both pathways generates a ladder without further intervention. This pair of steps may be repeated a controlled but unlimited number of times with drying between each step in order to generate a ladder of any desired length. Finally, the processed peptide is analyzed by MS, wherein the sequence of the peptide is deduced from the mass ladder. The ratio of cleavage to regeneration permits the distinction of isoleucine and leucine.

Abstract: A method and a device for the analysis of 5-hydroxyindoles or catecholamines with high sensitivity. New chemiluminescence labeling agents, 6-aminomethylphthalhydrazide or 1,2-bis(phthalhydrazino)ethylenediamine, are reacted with 5-hydroxyindoles or catecholamines to form their stable derivatives. The derivatives emit strong luminescence in the presence of an oxidizing agent. In the chemiluminescence detection method, there is extremely low background noise and thus the method enables analysis with high sensitivity.

Abstract: Novel alkoxythiocarbonylimidazoles provide new reagents for the N-terminal sequencing of small polypeptide samples. These reagents form an alkoxy thiourea derivative which is cleaved with acid to remove the N-terminal amino acid as a stable thiazolinone which does not rearrange to a thiohydantoin. This thiazolinone may be derivatized to provide a detectable group such as a fluorescent group or ionizable group detectable by mass spectrometry.

Abstract: Methods of producing biopolymer ladders and their use to obtain structural information about the biopolymer. The ladders are produced by setting up catalytic cleavage and terminating reactions at the end of biopolymer molecules. The terminating reactions terminate cleavage of a percentage of the biopolymer molecules at each round of cleavage.

Abstract: A method estimating a property or parameter of a nucleic acid material, which property or parameter is one to which the electrical conductivity of the nucleic acid material is related, comprises measuring the electrical conductivity of the nucleic acid material, and estimating from the measurement the property or parameter of the material by reference to a predetermined relationship between electrical conductivity and said property or parameter. This is based on the discovery that there are certain important properties of nucleic acids, the quantitative determination of which is frequently desirable, which can be assessed by measurement of the conductivity of a solution of the nucleic acid or acids. Changes in such properties may be reflected in corresponding changes of electrical conductivity. The concentration of nucleic acid in solution and the molecular weight of a species of nucleic acid are examples of important property which may be determined.

Abstract: The invention encompasses methods of detecting and/or quantifying .epsilon.(.gamma.-glutamyl)lysine isodipeptide by catalytically releasing lysine, then measuring free lysine.