CN1139810C

CN1139810C - Method and test paper of smei-quantitative determination of morphine by monoclone antibody immune chromatography

Info

Publication number: CN1139810C
Application number: CNB98112724XA
Authority: CN
Inventors: 慧张; 张慧; 唐江萍; 刘跃曾; 樊旭辉
Original assignee: CHANGSHA-CITY MENTAL DISEASES HOSPITAL
Current assignee: CHANGSHA-CITY MENTAL DISEASES HOSPITAL
Priority date: 1998-11-10
Filing date: 1998-11-10
Publication date: 2004-02-25
Anticipated expiration: 2018-11-10
Also published as: CN1259668A

Abstract

The present invention relates to a method and test paper for detecting morphine semi-quantitatively in a one-step immune chromatography method, which has the technical scheme that morphine in liquid to be detected and morphine marked on the surface of colloidal gold are competitively combined with morphine monoclonal antibodies accurately and quantitatively dropped on a chromatography film in a horizontal strip form; the content of the morphine in the liquid to be detected is determined by the number of stripes displayed on the chromatography film; the more the stripes are, the higher the content of the morphine in the liquid to be detected is; the bottom plate of the test paper and an immune chromatography complex film are sequentially superposed; colloidal gold is positioned at one end of the bottom plate and is in contact with the complex film; the chromatography film has good adsorption force to protein and has good wettability; the chromatography film is made of high molecular materials having a certain micro pore diameter.

Description

The method of smei-quantitative determination of morphine by monoclone antibody immune chromatography and test paper

Technical field: the present invention discloses a kind of method and test paper of smei-quantitative determination of morphine by monoclone antibody immune chromatography.

Background technology: the detection method to morphine in drug addict's body fluid can be divided three major types, i.e. chemical colour reaction analysis (as AC method etc.), stratographic analysis (as thin-layer chromatography, gas-matter coupling etc.) and immunoassay (as radioimmunology, immunochromatographic method etc.).The immunochromatography single stage method is to be set up by American scholar the early 1990s, and this method need not handled with its sample, and easy to be quick, characteristics such as suitable on-the-spot detection bringing into play very important effect in the drugs testing, but this method can only be qualitative, can not be quantitative; And its quality control system is another set of antigen-antibody, can not reflect the response situation of detection system truly; In addition, its reaction pattern shows as the positive and does not develop the color at detection zone, and negative for developing the color, this kind reaction pattern does not meet the thinking set of common result of determination.

Summary of the invention: the object of the invention is to overcome above-mentioned technical deficiency and a kind of high specificity is provided, and is highly sensitive, simple in structure, easy for operation, tentatively the method and the test paper of the smei-quantitative determination of morphine by monoclone antibody immune chromatography of detection by quantitative.

Fundamental purpose of the present invention is achieved in that by morphine in the liquid to be checked and the morphine that is marked at the collaurum surface and at least two accurate quantifications and is the horizontal stripe shape and selects the morphine monoclonal antibody band that schedules on the immunochromatography composite membrane competitive the combination taken place, by being shown in the striped quantity on the immunochromatography composite membrane, to determine the content of morphine in the liquid to be checked, fringe number is many more, show that morphine content is high more in the liquid to be checked, thereby finish the half-quantitative detection of morphine.

Another object of the present invention is achieved in that test paper comprises immunochromatography composite membrane, antigenic mark collaurum, base plate, thieving paper, base plate, immunochromatography composite membrane are stacked successively, one end of collaurum place base plate contacts with the immunochromatography composite membrane, the immunochromatography composite membrane is selected the morphine monoclonal antibody band that fixed at least two accurate quantifications are the horizontal stripe shape by protein being had good adsorption power and infiltrating micropore diameter macromolecular material is made on the immunochromatography composite membrane.

Embodiment: embodiments of the invention are summarized as follows:

One, the preparation of mark collaurum: get 1% gold chloride 1ml and be diluted to 100ml, add a certain amount of reductive agent under the state boiling, make collaurum with DDW.Be with morphine-6-O-half succinyl-bovine serum albumin(BSA) (Mor-BSA) collaurum to be carried out mark under the condition of 7-8.2 at pH value, carry out purifying with supercentrifugal process again, the accurate quantification freeze drying of instiling on glass fibre is standby.

Two, the special-purpose polymer compound film preparation of immunochromatography

At first make transparent backing film with a kind of macromolecular material, it is strong to protein bound power to cover one deck on backing film, the chromatographic film with certain pore size that wetting property is good.The effect of backing film is to stop organic solvent in the viscose binder to the destruction of protein on the chromatographic film.

Three, the point sample method of chromatographic film

With certain density morphine monoclonal antibody accurate quantification, be horizontal stripe shape point on chromatographic film with a determining deviation, use for detecting.

Four, the key technical indexes

1 specificity: only morphine class material is reacted;

2, sensitivity: 0.3 microgram morphine/milliliter sample;

3, sxemiquantitative scope: 〉=0.3 microgram～＞1.2 micrograms;

4, quick: observations in 10 minutes;

5, easy: anyone by specification can be operated;

6, be not subjected to such environmental effects.

Five, the application of test paper

1, to the detection of negative sample: three urine to be measured are splashed into well, keep flat observations after 5 minutes, have only a colour band negative.

2, the detection of positive sample: three urine to be measured are splashed into well, keep flat observations after 5 minutes, two above colour bands are arranged, and positive (〉=0.3 microgram/ml), three colour bands are about 0.8 microgram/ml, and four colour bands are 〉=1.2 micrograms/ml.

3, cross reaction: test paper and heroin and codeine have cross reaction, and intensity is similar to morphine; With methadone, naloxone, buprenorphine, meperidine, dihydroetorphine, fentanyl, diphenoxylate, cocaine, amphetamine, eastern Liang if alkali, ephedrine, haloperole, chlorpromazine, bucinnazine, fenazil, sodium phenobarbital, stable, lofexidine, methaqualone, somedon, clonidine, artane, caffeine, carbetapentane citrate, alprazolam, the equal no cross reaction of Clozapine.

Six, test paper reaction principle

Freeze and on collaurum, be marked with morphine antigen, be fixed with the morphine monoclonal antibody on the polymer compound film.When in the tested urine during no morphine, urine dilution antigenic mark collaurum also promotes its swimming on composite membrane, and when running into article one morphine monoclonal antibody, the morphine antigen on the collaurum will react fully with it, forms a macroscopic lilac band.When having morphine to exist in the tested urine, morphine in the urine will combine with article one morphine monoclonal antibody competitively with the morphine on the collaurum, make the collaurum of the anti-note of morphine not tackled by article one monoclonal antibody fully, can only continue swimming forward, show two colour bands with the reaction of second monoclonal antibody band.In like manner, morphine content is many in the urine, and colour developing tape number is also just many more, thereby can carry out half-quantitative detection to morphine.

In sum, high specificity of the present invention, highly sensitive, simple in structure, easy for operation, can effectively detect the roughly content of drugs in the liquid to be detected, be adapted to the instrument that procuratorial, judicial and public security organ, drug addiction treatment mechanism, hospital, employing unit, drug addict family members etc. detect as drugs.

Claims

1, a kind of method of immunochromatography smei-quantitative determination of morphine, the morphine that it is characterized in that morphine in the liquid to be checked and be marked at the collaurum surface and at least two accurate quantifications also are the horizontal stripe shape and select the morphine monoclonal antibody band that schedules on the immunochromatography composite membrane competitive the combination taken place, by being shown in the striped quantity on the immunochromatography composite membrane, to determine the content of morphine in the liquid to be checked, fringe number is many more, show that morphine content is high more in the liquid to be checked, thereby finish the half-quantitative detection of morphine.

2, a kind of test paper of immunochromatography smei-quantitative determination of morphine, comprise immunochromatography composite membrane, antigenic mark collaurum, base plate, thieving paper, base plate, immunochromatography composite membrane are stacked successively, one end of collaurum place base plate contacts with the immunochromatography composite membrane, protein is had good adsorption power to the immunochromatography composite membrane and infiltrating micropore diameter macromolecular material is made, it is characterized in that, on the immunochromatography composite membrane, select the morphine monoclonal antibody band that fixed at least two accurate quantifications are the horizontal stripe shape.