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Liquid-phase galactose oxidase-schiff's assay

A technology of oxidants and sugars, applied in biochemical equipment and methods, microbiological determination/inspection, measuring devices, etc., can solve the problems of reducing the background of non-specific reaction products, eliminate the uncertainty of false negatives, and be clinically beneficial Effect of performance, report time reduction

Inactive Publication Date: 2008-11-12
PREMD INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The prerequisite of sample fixation prior to GOS treatment has several drawbacks: 1) a drying phase to fix the clinical samples 2) rinses after galactose oxidase (GO) incubation and greater volume of washes after incubation with Schiff reagent to Reduce the background generated by non-specific reaction products, while removing excess enzyme and chromogen respectively 3) Manual processing of adsorbed samples 4) Subjective visual interpretation of results
Another drawback of the prior art is that, in order to detect glycoproteins and verify the presence of the sample on the solid phase, the GOS procedure is followed by the use of periodic acid-Schiff reagents

Method used

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  • Liquid-phase galactose oxidase-schiff's assay
  • Liquid-phase galactose oxidase-schiff's assay

Examples

Experimental program
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Effect test

Embodiment 1

[0055] Example 1: Rectal Mucus

[0056] Rectal mucus was obtained by digital examination with a lubricated, gloved finger. In order to evaluate the reactivity of the above-mentioned mucus with GOS, it is first necessary to immobilize the mucus on a water-insoluble substrate (such as a membrane filter, glass slide). For liquid-phase GOS assays, mucus is recovered from the glove with the aid of a solubilizing agent, preferably in small amounts to minimize sample dilution. Alternatively, mucus samples can be collected onto swabs (cotton, polyester, polyamide, foam, alginate), extracted, and then tested with GOS in solution. Calcium alginate constituents are particularly suitable for sample recovery due to the solvation of the test fiber in some reagents (sodium citrate, glycerol phosphate, sodium polymetaphosphate, EGTA or EDTA) to form clear gels or solutions . Mucus released into the gel / solution can be determined with the following GOS:

[0057] 1) Incubate dissolved algin...

Embodiment 2

[0060] Example 2: Saliva and lung sputum

[0061] Saliva is collected in a cup, which is fluid sufficient for aspiration without solubilization or mucolytic agent pretreatment. Centrifuge prior to aspiration analysis to remove host buccal and bacterial cells from saliva. Saliva obtained from coughed up phlegm can be centrifuged. Saliva can be directly treated with GOS.

[0062] Sputum is a thick, gelatinous respiratory secretion that, in addition to normal and abnormal lung cells, contains mucin macromolecules (high molecular weight glycoproteins), bacterial polysaccharides and genetic material, host leukocyte DNA and actin filaments. Sputum is often coughed up with saliva and can be separated by centrifugation. Typically, sputum is collected after deep inspiration and vigorous coughing, but induction with hypertonic saline (eg, ≥3% NaCl) may be required. The consistency of sputum renders it immiscible with aqueous reagents. This precludes direct reaction with GOS unless...

Embodiment 3

[0069] Example 3: Nipple discharge

[0070] Nipple discharge (NAF) may be clear, slightly cloudy and / or discolored. It facilitates imbibition, so NAF can be tested directly with the GOS procedure without disulfide reducing agent pretreatment. In general, the method is essentially the same as for rectal mucus, saliva, and lung sputum, with adjustments for sample and reagent volumes.

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Abstract

The invention provides an improved method for detecting cancer or a precancerous condition in a sample using an oxidation agent, such as galactose oxidase, and an aldehyde detection agent, such as Schiffs reagent that does not require the sample to be immobilized onto a solid support. The invention also provides kits comprising the components necessary for carrying out the methods of the invention.

Description

technical field [0001] The present invention relates to an improved assay for the detection of cancer or precancerous lesions using oxidizing agents and aldehyde detection reagents such as galactose oxidase and Schiff's reagent. The invention also relates to kits comprising the parts necessary to carry out the methods of the invention. technical background [0002] The galactose oxidase Schiff (GOS) assay is used to detect carbohydrate markers (such as D-galactose [Gal], or N-acetyl-D-galactosamine [ GalNAc], or D-Gal-β-[1→3]-N-acetyl-D-galactosamine [GalGalNAc] disaccharide [T or Thomsen-Friedenreich (TF) antigen]). These markers may be found in rectal mucus (colon cancer), saliva or sputum (oral cancer, lung cancer), nipple discharge (breast cancer), other mucous discharge (vaginal fluid [cervical cancer, uterine cancer, endometrial cancer] ]), semen [prostate cancer]) and blood (various cancers). Shamsuddin (US Pat. No. 5,348,860) discloses a method for detecting marke...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/26G01N33/50G01N33/574
CPCG01N33/57469G01N33/64G01N2333/904G01N33/5091G01N2400/38
Inventor C·卡特M·伊夫利R·扎瓦蒂斯基
Owner PREMD INC
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