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Analytical method, kit, and apparatus

a technology of kit and kit, applied in the field of assay method, can solve the problems of loss of treatment timing and lack of inclusion

Inactive Publication Date: 2001-07-05
NISSUI PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0037] As a reagent component to be used in accordance with the present invention, use may be made of "marker-labeled ligand" produced by binding a marker to a first ligand, and "bond element-labeled ligand" produced by binding a bond element consisting of nucleic acids with a predetermined base sequence, depending on the analyte species, to a second ligand. Separately from the developing element, the reagent component can compose a kit to be used in combination with the developing element. Additionally, the reagent component may satisfactorily be retained at a dry state in the sealing zone of the developing element.

Problems solved by technology

However, one laboratory reagent can assay or detect one item in most cases in the prior art, so the sample volume drawn from a patient is increased in proportion to the number of tests, which works as one of physical burdens over the patient.
Therefore, such device works as one cause to lose the timing for treatment because the doctor cannot make a decision instantly.
However, the publication does not include any description about the simultaneous assay or detection of one or more species of biological substances such as antigen or antibody.

Method used

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  • Analytical method, kit, and apparatus
  • Analytical method, kit, and apparatus
  • Analytical method, kit, and apparatus

Examples

Experimental program
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example 1

[0085] Process 1: Preparation of biotinylated oligonucleotide

[0086] As shown below, oligonucleotides each having amino acid at 5' terminus were individually generated synthetically by using a DNA synthesizer, 391 A, manufactured by Perkin Elmer, Co.

[0087] Amino group--GAA TTC CCG GGG ATC CGT CG

[0088] (referred to as pair 1+ hereinafter)

[0089] Amino group--CGA CGG ATC CCC GGG AAT TTC

[0090] (referred to as pair 1- hereinafter)

[0091] Amino group--AAC GGA ATC TAA TCA GGA GG

[0092] (referred to as pair 8+ hereinafter)

[0093] Amino group--CCT CCT GAT TAG ATT CCG TT

[0094] (referred to as pair 8- hereinafter).

[0095] These oligonucleotides each of 300 nmol were dissolved in 0.1 M MOPS buffer, pH 7.0 containing 1 mM EDTA, to which was added N-hydroxysuccinimide-biotin (30 .mu.mol; NHS-Biotin manufactured by Pierce, Co.) dissolved in N',N'-dimethylformamide (referred to as DMF hereinbelow), for reaction at 37.degree.C. for one hour. After the reaction, biotinylated oligonucleotides were separate...

example 2

[0115] Process 1: Preparation of (pair 1+)-labeled avidin and (pair 1-)-labeled avidin

[0116] The biotin-labeled pair 1+ (112 nmol) prepared at the process 1 in Example 1 was reacted with avidin (1.52 mg) in MPBS (0.7 ml) at 37.degree.C. for 3 hours, and subsequently, the mixture was loaded to Ultrogel AcA44 resin manufactured by IBF biotechniqes, preliminarily equilibrated with PBS, to recover pair 1+ labeled avidin. By the same method, additionally, pair 1+ labeled avidin was recovered.

[0117] Process 2: Preparation of oligonucleotide-avidin matrix-bound membrane

[0118] Avidin (10 mg) was dissolved in phosphate buffered physiological saline (manufactured by Nissui Pharmaceuticals, Co. PBS (-)),and then, membrane cut into a piece of 5.times.10 cm (SPHF membrane, manufactured by Millipore, Co.) was immersed in the solution at ambient temperature for one hour and was then rinsed with distilled water. After rinsing, the membrane piece was dried in air. By using a soft pen (manufactured b...

example 3

[0121] Process 1: Preparation of (pair 1-)-bound SPHF membrane

[0122] By using a soft pen (manufactured by Platinum Fountain Pen Co.) impregnated with 2 mg / ml (pair 1-)-labeled BSA as prepared at the process 4 in Example 1, a line was drawn vertically to the 5-cm side of a membrane cut into a size of 5.times.10 cm (SPHF membrane; manufactured by Millipore, Co.), to divide the side in halves, to bind the BSA through physical adsorption to the membrane. After drying in air, blocking by means of a blocking agent (Block Ace; manufactured by Snow Brand Milk Products, Co.) was effected at ambient temperature for 30 minutes, and subsequently, the resulting membrane was rinsed with distilled water and dried in air, to prepare (pair 1-)-bound SPHF membrane. After drying the membrane in air, the membrane was cut into a piece of a 0.5-cm width and a 5-cm length, and at one end of the piece was fixed GF with a staple, and storage under dry conditions.

[0123] Process 2: Capture of sandwich immunoc...

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Abstract

Providing an assay method capable of simultaneously determining the presence or absence of one or more species of biological substances or assaying the amounts thereof with a single assay device, a kit therefor and an assay device thereof. The amount thereof or the presence thereof is detected, by putting a liquid sample containing one or more species of analytes in contact to a reagent including one or more species of marker-labeled ligands and one or more species of nucleic acid-labeled ligands, to generate one or more species of complexes, developing the generated one or more species of complexes through capillary phenomenon in developing element 11 in a sheet form, capturing the complexes through complementary nucleic acid binding onto anti-bond elements consisting of nucleic acids on detection zones 15, 16 and 17 formed depending on each of one or more species of nucleic acids immobilized on the detection zone 14, thereby capturing a complex depending on the analyte species, through the complementary binding between the anti-bond element and the bond element, to form an independent band and to assay the amount or the presence on the detection part.

Description

[0001] The present invention relates to an assay method for assaying an analyte as a biological assay subject or for detecting the presence or absence thereof, which is useful for simple clinical diagnosis, and a kit and an assay device to be used for the method; more specifically, the present invention relates to an assay method for assaying a great number of combinations of one or more species of analytes contained in a fluid sample or the presence or absence thereof, and a kit and an assay device therefor.[0002] For determining the disease affecting a patient at a laboratory test, several types of laboratory test results should collectively be examined. In general, patients should undergo several types of such tests for appropriate diagnosis and therapeutic treatment. However, one laboratory reagent can assay or detect one item in most cases in the prior art, so the sample volume drawn from a patient is increased in proportion to the number of tests, which works as one of physica...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C12M1/00C12Q1/68G01N33/543G01N33/53G01N33/547
CPCG01N33/54353G01N33/54386
Inventor OKU, YUICHITANAKA, YOSHITATSUOTSUKA, YOKO
Owner NISSUI PHARMA
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