CN2869864Y - Colloid gold test-paper for quick detecting benomyl residue - Google Patents

Abstract

A colloidal gold test paper used for the rapid detecting of benomyl residues, composing a base plate, a water absorption board, a pyroxylin membrane, a monoclonal antibody gold label mat, and a specimen imbibition layer. The middle of the base plate is a pyroxylin membrane on which is provided with benomyl synthetic immunogen test line and a multi- clone antibody control line; one end tip of the base plate is provided with a water absorption board, and the other end tip is provided with a specimen imbibition layer; the two ends of the pyroxylin membrane are separately overlapped for connection each other with the water absorption board and the monoclonal antibody gold label mat; the top of the monoclonal antibody gold label mat is pressed with a specimen imbibition layer; the test paper is made by means of a colloidal gold immunity competition method; the immunity colloidal gold method is used to detect if the benomyl level of residue in garden stuff is overproof. The method has the features of speediness, sensitivity, simplicity of operation, low cost, no necessity of detecting by professional, real attainment of integrating the complicated principle with the simplicity of operation organically, as well as convenience of preservation and long period of validity.

Description

The colloid gold test paper that the fast detecting benomyl is residual

Technical field

The present invention relates to a kind of vegetables and raw grain based food Detecting Pesticide test paper and detection method thereof of being used for, belong to the colloid gold immune detection range.

Background technology

Because the vegetable growth phase is short and disease and pest is comparatively serious, thereby, in the growing vegetables process, usually need the continuous several times dispenser.And many vegetables need repeatedly be gathered, gather and the time interval of dispenser short, cause when gathering the persticide residue in the vegetables often higher, be easy to surpass state specified standards.And persticide residue exceeds standard and can have a strong impact on level of human health in the vegetables, has become the food pollution source that can not be ignored.Government and each functional department extremely pay close attention to the residues of pesticides problem of food.But because the resistance to the action of a drug of the toxicity of agricultural chemicals and insect forms a vicious cycle, forbid thoroughly comprehensively that therefore the use of high highly toxic pesticide will have long process, even in using, low-toxin farm chemicals comprehensively, the pernicious poisoning of agricultural chemicals still takes place.Therefore new requirement and higher standard have all been proposed all many-sides such as the object of analyzing and testing, kind, quantity, scope, index.

Benomyl (1-n-butyl amine base formyl-methyl 2-benzimidazolecarbamate), the multiple diseases that can be used for preventing and treating crops such as vegetables, fruit tree, rice wheat is commonly used for seed dressing, spraying and soil treatment, except that protection and therapeutic action are arranged, also has acaricidal action.This agricultural chemicals is the product that E.I.Du Pont Company produced more than 30 year, and owing to reasons such as toxicity, lawsuit is continuous, and E.I.Du Pont Company has to stop to produce in calendar year 2001.Benomyl can make allergic, takes with the gavage heavy dose in animal used as test but does not contact in recipe, can be observed teratogenic effect.Suck with oral and contact the spermatogenesis activity that can reduce animal used as test.In the pesticide residue joint conference various agricultural product and food have been advised maximum residual level in food that FAO/WHO holds and the environment, nineteen eighty-three, JMPR has determined that acceptable daily intake (ADI) is 0～0.02mg/kg/b.w..The U.S. had carried out clearly regulation to the maximum residue limit of benomyl in the multiple kinds of crops in 1998, and the residual scope of limiting the quantity of of regulation benomyl is 0.2mg/kg～15mg/kg in this standard.China limits the quantity of to benomyl in the crops residual at present and does not make clearly regulation as yet, so the standard of formulating with reference to the U.S..

At present, the analytical approach of detection residues of pesticides mainly is dependence, vapor-phase chromatography (GC) or gas chromatography and mass spectrum (MS) coupling method (GC/MS)." enzyme inhibition rate method+spectrophotometric method " has been listed in national proposed standard (GB/T 5009.199-2003).This method is quick, sensitive, easy and simple to handle and with low cost, can realize quick primary dcreening operation, but the restriction of examined scope and precision.And vapor-phase chromatography (GC) can realize the quantitative test of precision but speed is slower, and it is higher to detect cost.In a word, these method ubiquities the sample pretreatment process complexity, sensitivity be subjected to sample purification, step such as concentrate influence very big, consuming time, checkout equipment costliness, and require to have technical professional and long analytical cycle.

Therefore, utilize the competition law principle, on the basis of traditional immunodetection, introduced the collaurum Fast Detection Technique, develop benomyl collaurum competition law and detect test paper.This method has fast, sensitive, easy and simple to handle, cost is low, need not the professional detects, and really reaches the organic unity of complicated principle and ease of Use, have simultaneously preserve convenient, the characteristics that the term of validity is long.

Summary of the invention

The present invention is directed to some problems of above-mentioned existence, provide a kind of being applicable to detect the collaurum mensuration test paper whether benomyl exceeds standard.

For solving the problems of the technologies described above, the present invention is achieved by the following technical solutions:

At benomyl residue detection test paper base plate middle part is nitrocellulose filter, a test wire and a sheep anti mouse polyclonal antibody control line are arranged on the nitrocellulose filter, in base plate one end termination is water accepting layer, other end termination is the sample liquid-adsorption layer, the nitrocellulose filter two ends overlap mutually with benomyl monoclonal antibody gold mark pad with water accepting layer respectively and are connected (the overlapping coupling part is in 1-2 millimeter scope), are pressed with the sample liquid-adsorption layer on benomyl monoclonal antibody gold mark pad.Combined method about base plate, nitrocellulose filter, thieving paper, collaurum mark pad is carried out by patent CN 2741052Y.

The control line of benomyl residue detection test paper is formed by sheep anti mouse polyclonal antibody bag, and test wire is by benomyl-synthetic immunogen bag quilt.The sample liquid-adsorption layer end that detects the residual colloid gold test paper of benomyl is put into Juice, because the capillarity sample will move along test strips water accepting layer end, when moving to benomyl monoclonal antibody gold mark pad, benomyl in the sample and benomyl monoclonal antibody gold mark probe generation specific bond, when moving to when being fixed with benomyl-synthetic immunogen and connecting the thing test wire, lose it and combine owing to benomyl in the antibody in the benomyl monoclonal antibody gold mark pad and the sample preferentially is combined into compound with benomyl-synthetic immunogen, therefore its collaurum can not be stranded on the test wire, the test wire place does not have red line to show, promptly has only a red control line positive; If do not have benomyl in the opposite sample, antibody in the benomyl monoclonal antibody gold mark pad moves on the test wire, the benomyl monoclonal antibody will connect thing generation specific bond with benomyl-synthetic immunogen, collaurum is stranded on the test wire, promptly two red line are negative, immune competition law principle that Here it is.Benomyl content just is inversely proportional in the shade of test wire and the sample.Carry out the transition to feminine gender from the positive, because the benomyl content difference of surveying, the color of test wire is also different, forms a red color gradient difference, detects the OD value with this principle, thereby infers the amount of actual benomyl that test wire reacts.

It detects more than the benomyl setting detected level to make test paper with competition law, and it is positive that a red line appears in the view window place; It is negative to set the two red lines of the following appearance of detected level; Be that fuzzy hacures are boundary value at the test wire place when setting detected level, thereby draw judgement.

When moving to sheep anti mouse polyclonal antibody control line, no matter have or not benomyl in the sample, the gold mark probe of mark all can combine delay with the sheep anti mouse polyclonal antibody that has configured, and it is red that control line is shown.Therefore control line does not have colour band and produces that then the representative operation is wrong, during detection the sample liquid level surpass the MAX line or test paper expired.

Because adopt technique scheme, the residual colloid gold test paper of a kind of fast detecting benomyl provided by the present invention has such beneficial effect, promptly high specificity is highly sensitive, easily stores, need not the technical skill personnel operation, and readability as a result.

Description of drawings

Fig. 1 is the main TV structure figure of the residual colloid gold test paper of a kind of fast detecting benomyl of the present invention.

Fig. 2 is the side-looking structural drawing of invention figure one.

Fig. 3 shows positive findings figure for detecting.

Fig. 4 shows negative findings figure for detecting.

Among the figure 1, water sucting plate, 2, nitrocellulose filter, 3, sheep anti mouse polyclonal antibody control line, 4, test wire, 5, monoclonal antibody gold mark pad, 6, the sample liquid-adsorption layer, 7, base plate, 8, the MAX line.

Specific embodiment

1. according to the molecular structure of benomyl, infer the antigen determining area in its molecule, select for use the benomyl pesticide original medicine synthetic a kind of close with the benomyl molecular structure and have carboxyl or amino etc. and be easy to the be connected molecular structure (haptens) of group of protein macromolecule through chemical reaction.The haptens micromolecule is connected with protein macromolecule makes the benomyl immunizing antigen.

2. MONOCLONAL ANTIBODIES SPECIFIC FOR

(1) with synthetic immunogen immune BALB/c mouse.Set up the knurl strain with the SP2/0 Fusion of Cells and screen, get oncocyte and be injected in the BALB/c mouse abdominal cavity, make it produce ascites.

(2) extract the screening of mouse ascites purifying, the monoclonal antibody that acquisition can combine with the benomyl molecule is used for colloid gold label.

The preparation of collaurum and with the combining of monoclonal antibody

(1) get distilled water and add an amount of gold chloride magnetic agitation and be warmed to 85～95 ℃, add an amount of citrate three sodium and continue heated and stirred to seething with excitement 3～10 minutes, it is standby to keep in Dark Place after the cooling.

(2) the collaurum liquid of benomyl labeling of monoclonal antibody, on the adsorbing fiber material, dry back is standby.

4. film-making machine system film: utilize computer control transmission speed, guarantee that the antibody amount of bag quilt on the per unit film equates.

5. test strips combination: see Figure of description, method is with patent CN 2741052Y.

6. using method: the sample liquid-adsorption layer end of test paper is put into vegetables soak juice (liquid level must not surpass MAX line chart 8), take out test paper after 1 minute and keep flat, because kapillary and syphonic effect sample will move observations in the time of 5 minutes along test strips water accepting layer end.

7. the result judges: detect more than the benomyl concentration 0.2mg/kg, it is positive that a red line appears in the view window place; 0.2mg/kg the two red lines of following appearance are negative; 0.2mg/kg the time be that fuzzy hacures are boundary value at the test wire place, this boundary value is the vegetable pesticide residue examination criteria 0.2mg/kg of country's promulgation.Thereby draw judgement.

Claims (6)

1. colloid gold test paper that the fast detecting benomyl is residual, it is characterized in that by base plate (7), water sucting plate (1), nitrocellulose filter (2), benomyl monoclonal antibody gold mark pad (5), sample liquid-adsorption layer (6) form, MAX line (8); The base plate middle part is a nitrocellulose filter, a test wire (4) and a polyclonal antibody control line (3) are arranged on the nitrocellulose filter, base plate one end termination is a water sucting plate, other end termination is the sample liquid-adsorption layer, the nitrocellulose filter two ends overlap mutually with monoclonal antibody gold mark pad with water sucting plate respectively and are connected, and are pressed with the sample liquid-adsorption layer on monoclonal antibody gold mark pad.

2. the residual colloid gold test paper of a kind of fast detecting benomyl according to claim 1 is characterized in that control line is to be made into by sheep anti mouse polyclonal antibody bag.

3. the according to claim a kind of residual colloid gold test paper of fast detecting benomyl is characterized in that test wire is to be made into by benomyl synthetic immunogen bag.

4. according to claim 1 or the residual colloid gold test paper of 3 described a kind of fast detecting benomyls, it is characterized in that the object that is detected is the potpourri that the former cartridge bag of benomyl is drawn together the benomyl preparation.

5. the residual colloid gold test paper of a kind of fast detecting benomyl according to claim 1 is characterized in that gold mark pad is as colloid gold label by the benomyl monoclonal antibody.

6. according to the residual colloid gold test paper of the described a kind of fast detecting benomyl of claim 1, it is benomyl 0.1～15mg/kg that the detected value that it is characterized in that test wire is set at limit of identification.

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