CN2511976Y - Indicator paper for identifying cancer from hyperplasia - Google Patents

Abstract

The utility model relates to an indicator paper for identifying cancer from hyperplasia using one-step method detection technology with monoclonal antibody and colloidal gold immunity chromatography and a method of determining ratio between dissociating PSA and total PSA in whole blood or serum of human body by the indicator paper. The indicator paper invented is provided with a detection line; the data of the detection line sets that the ratio between dissociating PSA and total PSA is 10 percent which is used as a dividing value of chromogenesis and non-chromogenesis. Whether the detection line is chromogenic or not is a mark of the ratio between dissociating PSA and total PSA in whole blood or serum of human body; when the indicator paper is used, no any reagent is required to add except adding tested samples; no washing and no other separation procedures are required, thereby being simple, rapid and convenient in operation, intuitionistic in detecting results, easy in judgment, and the indicator paper (II type) and the method of determining the ratio between dissociating PSA and total PSA in whole blood or serum of human body by the indicator paper can identify prostate cancer and in particular early prostate cancer or prostate hyperplasia.

Description

A kind of examination prostate cancer and hyperplasia of prostate test paper

Technical field: the utility model relates to a kind ofly to be made the test paper of screening prostate cancer and hyperplasia of prostate and uses this test paper to measure in human whole blood or the serum the free PSA and the total method of PSA ratio with monoclonal antibody and colloidal gold immunochromatographimethod single stage method detection technique.Belong to the articles for use field of detecting.

Background technology: prostate cancer is the common disease frequently-occurring disease of middle-aging male.According to statistics, in the male cancer patient, the incidence of disease of prostate cancer is the highest, is the tertiary cause of the death except that lung cancer and colorectal cancer.Prostate characteristic antigen PSA is that molecular weight is the highly tissue-specific glycoprotein of having of 34KD, except being distributed in normal prostata tissue, hyperplastic prostate tissue, malignant prostate tissue, prostate metastatic carcinoma and prostate juice, and refining China and foreign countries, PSA is not present in other any normal structure, is not present in as in other cancers such as breast cancer, lung cancer, carcinoma of the colon and rectum, cancer of the stomach, cancer of pancreas and thyroid cancer yet.For the patient male sex of normal health and non-prostatic lesion, the content of its whole blood or blood-serum P SA all is lower than 4.0ng/ml.When patient suffered from the adjacent reproduction urethral tissue inflammation of prostate cancer, hyperplasia of prostate (BPH) or other, remarkable rising can take place in whole blood or blood-serum P SA concentration, and therefore, PSA is the detection mark of effective prostate cancer so far.The PSA that detects in whole blood or the serum is the diagnosis of carrying out prostate cancer, monitors generation, the development of its illness, estimates the effect after its treatment, and the important indicator that detects the recurrence rate after its chemotherapy or the radiotherapy.The kit that can be used for the detection of PSA serum at present has radio-immunity (RIA) kit, enzyme mark (ELISA) immune reagent kit, photochemistry immune reagent kit and fluorescent mark immunity kit etc.All need be equipped with corresponding instrument when using these kits, and require to have the professional and technical personnel to operate and to carry out.During mensuration, determination step is many, and the time is long, and the expense height is restricted the use of these methods.Also there is the test strips of measuring blood-serum P SA to occur at present abroad, but its product can only show the situation that increases of PSA in the serum, and there is the gray area phenomenon in measuring, promptly when the total PSA of the serum in the working sample is between 4-10ng/ml, it is because prostate cancer, especially early prostate cancer still are other prostatic disorder causes as hypertrophy of the prostate etc. that the PSA that can't distinguish this moment raises.Therefore must further distinguish with this most reliable existing conclusive evidence standard of prostate biopsy again, and 25% the people of only having an appointment in the reality may be a prostate cancer, this patient of other 75% who just means that the total PSA of those serum is positioned at gray area also has to get rid of by prostate biopsy, and this bring the expense expenditure of unnecessary misery and great number can for undoubtedly these patients' body and mind.Simultaneously since at present the PSA clinical detection method of using all with serum as sample, so just making serum separate inconvenient user can't use, and the use of these assay methods is restricted.Multinomial result of study proves: the PSA value should be made the mensuration of free PS A and total PSA number percent in the gray area scope of 4.0-10.0ng/ml, can improve the sensitivity and the specificity of diagnosis so greatly, distinguish prostate cancer comparatively exactly, especially early prostate cancer and general prostatic disorders, as hyperplasia of prostate, avoid unnecessary biopsy.Research also shows: when free PS A is respectively between the 20-25% with the total ratio of PSA, between the 15-20%, between the 10-15% and between the 0-10% time, the ratio that may suffer from prostate cancer then is respectively 16%, 20%, 28% and 56%.International Federation of Clinical Chemistry in 1997 determines the general ratio of free PS A and total PSA is decided to be 10% for this reason.

Summary of the invention: the defective that the objective of the invention is to overcome prior art, invent a kind of examination prostate cancer and hyperplasia of prostate test paper that utilizes colloidal gold immunochromatographimethod single stage method principle and monoclonal antibody technique to make, a detection line is arranged in this test paper, detection line colour developing setting value is 10% as colour developing and the cut off value that does not develop the color for the ratio with free PS A and total PSA, and to the detection method of free PSA numerical value in human whole blood or the serum with the ratio of total PSA.Base need not other any reagent when detecting, do not need washing or other separation steps yet, can screen prostate cancer, especially the test paper of early prostate cancer and hyperplasia of prostate (II type) and use this detection paper human whole blood or serum in the method for free PSA numerical value and the ratio of total PSA, it detects accuracy rate and can reach more than 90%.Another object of the present invention is to avoid serum to separate trouble, can directly use whole blood to carry out the test paper of above-mentioned testing and examination prostate cancer and hyperplasia of prostate.

The utility model is finished by following manner: screen prostate cancer and hyperplasia of prostate test paper, include base plate, inhale sample film, the cellulose nitrate rete, collaurum film and water accepting layer, on base plate, be fixed with the cellulose nitrate rete, it is characterized in that being fixed with one section hydroscopic substance on cellulose nitrate rete top, constitute water accepting layer, cellulose nitrate rete middle part is sprayed with an anti-prostate characteristic antigen PSA monoclonal antibody, constitute detection line, between suction sample rete and cellulose nitrate rete, accompany the collaurum rete, the gold mark probe that contains useful another anti-PSA Monoclonal Antibody in the collaurum film is added to divide the blood film to constitute the whole blood test test paper into one deck on the suction sample film.

The detection of free PSA and total PSA ratio in human whole blood or the serum a: detection line is arranged in the test paper, detection line develops the color or does not develop the color, as measuring free PSA and total PSA ratio sign in human whole blood or the serum, may further comprise the steps during mensuration: A, test paper is inhaled the sample film end, and to insert in the test sample 0.5-1.0cm dark, keep 10-30 second, keep flat after taking out test paper, perhaps sample is directly dropped in test paper and inhale the sample film end, observe the detection line place after 5-30 minute and whether develop the color; B, when detection line does not develop the color, show in the sample that the content of free PSA surpasses 10%, the possibility of suffering from prostate cancer is less, can make prostate biopsy, pathological section etc. and further check; C, when detection line develops the color, show in the sample that the content of free PSA is less than or equal to 10%, the possibility of suffering from prostate cancer is bigger, should do prostate biopsy, pathological section etc. and further check; The ratio that the detection line data are set to free PS A and total PSA is 10%, as colour developing and the cut off value that does not develop the color.

Use the present invention to screen that the method for total PSA and free PS A ratio is in prostate cancer and hyperplasia of prostate detection paper human whole blood or the serum: test paper is inhaled 10-30 second in the sample film end insertion test sample, perhaps directly testing sample is dropped in and inhale sample film one end, then inhaling sample film (1) can make test paper possess the sample liquid of some, sample liquid is by the capillary action of nitrocellulose filter, sample liquid can be carried to the other end from test paper one end, PSA in the course of conveying in the sample liquid at first combines with anti-PSA gold mark probe generation specificity in the collaurum rete (2), continue transported to when reaching detection line (3), again with detection line in another anti-PSA monoclonal antibody specificity of forming Sanming City smelting formula combine and stop, so the detection line colour developing, detecting overall process need be with 5-30 minute.When measuring whole blood sample, be arranged in the branch blood film (7) inhaled on the sample film (1) and can be automatically the haemocyte of blood be separated with serum, to avoid the influence of haemocyte to test paper mensuration.

When total PSA numerical value is confirmed as 4-10ng/ml in detecting whole blood or serum, just need with free PSA in examination prostate cancer shown in the present and hyperplasia of prostate detection paper human whole blood or the serum and total PSA ratio, to determine that prostate cancer still is other prostatic disorders such as hypertrophy of the prostate, determine perhaps whether the experimenter needs to do biopsy or other further inspection.

When the present invention screens detection line (3) colour developing of prostate cancer and hyperplasia of prostate test paper, the content of free PSA is less than or equal to 10% in the interpret sample, it is bigger to show that the patient suffers from the possibility of prostate cancer, should do further prostate biopsy or check pathological section etc. and further check.

The utility model prostate cancer and hyperplasia of prostate are screened the test paper color development principle: utilize colloidal gold immunochromatographimethod single stage method principle and monoclonal antibody technique, the a certain zone that in advance the anti-PSA monoclonal antibody of certain numerical value is fixed on nitrocellulose filter constitutes detection line, and another anti-PSA monoclonal antibody that will combine with collaurum, promptly anti-PSA gold mark probe stationary is in another zone of collaurum rete, during mensuration test paper is inhaled sample film one end and insert 10-30 second in tested whole blood or the blood serum sample, perhaps directly testing sample is dropped in and inhale sample film one end, then inhaling sample film (1) can make test paper maintain the sample liquid of some, sample liquid can be by the capillary action of nitrocellulose filter, carry to the other end from test paper one end, PSA in course of conveying in the sample liquid at first with the collaurum rete in anti-PSA gold mark probe generation specific bond, when continuing transported to the detection line position on the nitrocellulose filter, combine with another anti-PSA monoclonal antibody specificity of forming Sanming City smelting formula in the detection line again, so stop, when contained free PS A in whole blood or the blood serum sample is less than or equal to the colour developing numerical value that sets with the ratio of total PSA, the detection line colour developing, by setting a detection line that detects numerical value, differentiate the ratio of contained free PS A in this whole blood or the blood serum sample and total PSA, further specify the possibility that the tested person suffers from prostate cancer, the branch blood film that sets in advance during whole blood assay can separate haemocyte in the blood and serum, avoids the influence of haemocyte to measuring.Whole mensuration needs 5-30 minute.

The utility model test paper in use, except that the needs sample, need not to add any reagent, need not wash yet, not need other separating step, simple to operate, quick, convenient, testing result is directly perceived, be easy to judge, and can screen prostate cancer, especially the method for the ratio of free PSA numerical value and total PSA in the test paper of early prostate cancer or hyperplasia of prostate (II type) and this detection paper human whole blood of use or the serum.

Description of drawings: describe below in conjunction with accompanying drawing.

Fig. 1 screens prostate cancer and hyperplasia of prostate test paper longitudinal sectional view for the utility model.

Fig. 2 is for after using detection paper whole blood shown in Figure 1 or blood serum sample, and detection line does not develop the color, and illustrate that free PSA in whole blood or the serum and total PSA ratio are greater than 10% user mode figure.

Fig. 3 is for after using detection paper whole blood shown in Figure 1 or blood serum sample, and the detection line colour developing illustrates that free PSA is less than or equal to 10% user mode figure with total PSA and ratio in whole blood or the serum.

Embodiment (test paper manufacturing): preparation the utility model is screened prostate cancer and hyperplasia of prostate test paper.Select self-produced (Kunming cloud mcroorganism technology company) for use or buy the anti-PSA monoclonal antibody of two kinds of different detection site of (as U.S. Sigma company etc.), use wherein a kind of Antibody Preparation gold mark probe, and in advance the anti-PSA antibody of a certain amount of another kind is sprayed on certain zone of nitrocellulose filter, form a detection line, choose long 100 millimeters, wide be 80 millimeters, thick be that 1 millimeter white PVC plastics are base plate, by double faced adhesive tape paste in the centre earlier length be 100 millimeters, wide be 30 millimeters the nitrocellulose filter that is sprayed with a detection line; Paste length again on the top of nitrocellulose filter and be 100 millimeters, wide be 30 millimeters hydroscopic substance (as filter paper etc.), the formation water accepting layer; The other end at nitrocellulose filter is pasted the collaurum rete, and it has the gold mark probe that contains with above-mentioned anti-PSA Monoclonal Antibody, pastes on the collaurum rete and inhales sample film, after stickup finishes, be cut into 80 millimeters long, the test strips of 2.5 mm wides, packing gets final product.If the whole blood test test paper of whole blood sample can be directly measured in preparation, need inhaling how stickup one decks divide the blood film on the sample rete.

Screen embodiment:

1, preparation PSA titer, wherein free PS A is 25% with the ratio of total PSA, the ratio of free PS A and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert sample liquid, the degree of depth is 0.5cm, keep taking out after 10 seconds, keep flat after 10 minutes and observe, no any colour band manifests on the test paper.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 10 minutes and observe, no any colour band manifests on the test paper.

2, preparation PSA titer, wherein free PS A is 10% with the ratio of total PSA, the ratio of free PS A and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert sample liquid, the degree of depth is 0.8cm, keep taking out after 25 seconds, keep flat after 10 minutes and observe, color band manifests on the test paper.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 10 minutes and observe, color band manifests on the test paper.

3, preparation PSA titer, wherein free PS A is 5% with the ratio of total PSA, the ratio of free PS A and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert sample liquid, the degree of depth is 0.6cm, keep taking out after 20 seconds, keep flat after 15 minutes and observe, color band manifests on the test paper.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 15 minutes and observe, color band manifests on the test paper.

4, always learning from else's experience, PSA measures, its value is that 4.2ng/ml patients blood or serum 0.5ml place the mensuration test tube, the ratio of free PSA and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert in whole blood or the serum, the degree of depth is 0.5cm, keeps taking out after 15 seconds, keeps flat after 10 minutes to observe, no colour band manifests, and the ratio that free PS A and total PSA in this blood serum sample are described is greater than 10%.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 10 minutes and observe, no colour band manifests, the ratio that free PS A and total PSA in this sample are described is greater than 10%.

5, always learning from else's experience, PSA measures, its value is that 5.5ng/ml patients blood or serum 0.5ml place the mensuration test tube, the ratio of free PSA and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert in whole blood or the serum, the degree of depth is 0.5cm, keeps taking out after 30 seconds, keeps flat after 8 minutes to observe, color band manifests, and the ratio that free PS A and total PSA in this sample are described is less than 10%.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 8 minutes and observe, color band manifests, the ratio that free PS A and total PSA in this sample are described is less than 10%.

6, always learning from else's experience, PSA measures, its value is that 6.1ng/ml patients blood or serum 0.5ml place the mensuration test tube, the ratio of free PSA and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert in whole blood or the serum, the degree of depth is 0.6cm, keeps taking out after 20 seconds, keeps flat after 5 minutes to observe, no colour band manifests, and the ratio that free PS A and total PSA in this sample are described is greater than 10%.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 5 minutes and observe, no colour band manifests, the ratio that free PS A and total PSA in this sample are described is greater than 10%.

7, always learning from else's experience, PSA measures, its value is that 9.1ng/ml patients blood or serum 0.5ml place the mensuration test tube, the ratio of free PSA and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert in whole blood or the serum, the degree of depth is 0.5cm, keeps taking out after 10 seconds, keeps flat after 10 minutes to observe, color band manifests, and the ratio that free PS A and total PSA in this sample are described is less than 10%.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 10 minutes and observe, color band manifests, the ratio that free PS A and total PSA in this sample are described is less than 10%.

8, always learning from else's experience, PSA measures, its value is that 9.6ng/ml patients blood or serum 1.5ml place the mensuration test tube, the ratio of free PSA and total PSA in the test strips working sample of method for preparing is taked in use, assay method is: (1), test strips is inhaled sample film one end insert in whole blood or the serum, the degree of depth is 0.5cm, keeps taking out after 15 seconds, keeps flat after 12 minutes to observe, no colour band manifests, and the ratio that free PS A and total PSA in this sample are described is greater than 10%.(2), above-mentioned sample liquid directly dropped in test paper inhale sample film one end, keep flat after 12 minutes and observe, no colour band manifests, the ratio that free PS A and total PSA in this sample are described is greater than 10%.

One, sensitivity is measured:

1, adopts the phosphate buffer preparation of PH7.4 to contain free PS A and total PSA ratio and be respectively 1%, 2.5%, 5%, 7.5%, 10% series standard liquid and blank, randomly draw the test strips that adopts method for preparing to above titer and blank replicate determination three times, when measuring free PS A and total PSA ratio and being respectively 1%, 2.5%, 5%, 7.5%, 10% titer, detection paper line (3) all should develop the color, and does not all develop the color in above mensuration empty contrast.

2, adopt the phosphate buffer preparation of PH7.4 to contain free PS A and total PSA ratio and be respectively 12.5%, 15%, 17.5%, 20%, 30% series standard liquid and blank, randomly draw the test strips that adopts method for preparing to above titer and blank replicate determination three times, mensuration contains free PS A and total PSA ratio when being 12.5%, 15%, 17.5%, 20%, 30% series standard liquid, detection paper line (3) does not all develop the color, and does not all develop the color in above mensuration empty contrast.

Two, specific assay: (PAP), albumin (Albumin), alpha-fetoprotein (AFP), carcinomebryonic antigen (CEA), glycoprotein (Glycoprotein), human IgG, cholerythrin (Bilirubin), CA19-9, CA125 are added in the serum that does not contain PSA, make that its concentration reaches 0,10,50,100 respectively, 200ng/ml, detect with test strips, its result is all negative, and above-mentioned material that may exist in cancer patient and the equal no cross reaction of this test paper are described.

Three, interference is measured: following material being added to the serum that does not contain PSA or contain free PS A and total PSA ratio is to test in 10% the serum.Interfering material comprises: acetaminophenol 20mg/dl, acetoacetate 1500mg/dl, acetone 1250mg/dl, acetylsalicylic acid 20mg/dl, albumin 1.4g/dl, ampicillin 40mg/dl, vitamin C 40mg/dl, biotin 30 μ g/dl, caffeine 40mg/dl, codeine 5 μ g/ml, corfisol 150ng/ml, creatinine 200mg/dl, DHEAS 10000ng/ml, estrone 25ng/ml, estradiol 25ng/ml, estriol 25ng/ml, ethanol 4000mg/dl, protoheme 30g/l, hedroxybulyric acid 100mg/dl, glucose 20g/l, oxalic acid 5000mg/dl, progesterone 50ng/ml, benzene Ba Bituo 10 μ g/ml, sallcysic acid 1500mg/dl, Si Kebabituo 10 μ g/ml, sodium carbonate 1500mg/dl, sodium chloride 5000mg/dl, tetracycline 40mg/dl, urea 3500mg/dl, uric acid 10mg/dl.Every group of test done three parts.The gained result is: all samples that does not contain PSA no matter whether contain interfering material, are negative findings; Total all free PS A and PSA ratio of containing is 10% blood serum sample, regardless of whether containing interfering material, is positive findings, illustrates that they all do not have obvious interference to the mensuration of PSA.

This patent screens prostate cancer and hyperplasia of prostate test paper and has that detection speed is fast, accuracy is high, safe and simple, be easy to use.During this detection paper, except that need add sample, need not to add any reagent, need not wash yet, do not need separation steps, simple to operation, testing result is easy to judge, and its testing result accuracy rate height is the prostatic cancer early diagnosis that uses of a kind of suitable medical institutions, health care department and family and individual and differentiate with hypertrophy of the prostate and the of new generation biological high-tech product of generaI investigation usefulness.

The present invention screen prostate cancer and hyperplasia of prostate test paper since can the human body whole blood or serum in the ratio of total PSA and free PS A, the patient that its value when detecting total PSA can be positioned at gray area does further differentiation, can be effectively with hypertrophy of the prostate patient and prostate cancer, especially the early stage patient of prostate cancer separates, efficient reaching more than 90%.Invention by this test paper, can improve prostate cancer, especially accuracy and recall rate that early stage prostate detects, can carry out in time and treatment targetedly patient thus, improve the cure rate of prostate cancer, reduce the mortality ratio of middle-aging male prostate cancer, and can exempt the unnecessary biopsy of part or other detection, reduce patient's misery, reduce testing cost, save detection time, be beneficial to and promote the use of, particularly the whole blood test test paper can be used for health screening more easily, family tests oneself and self-examination.

1-suction sample film; 2-collaurum film; 3-detection line; 4-water accepting layer; 5-base plate; 6-Nitrocellulose filter; 7-minute blood film.

Claims (3)

1, a kind of examination prostate cancer and hyperplasia of prostate test paper, include base plate (5), inhale sample film (1), nitrocellulose filter (6) layer, collaurum film (2) and water accepting layer (4), on base plate (5), be fixed with nitrocellulose filter (6) layer, it is characterized in that being fixed with one section hydroscopic substance on cellulose nitrate rete (6) top, constitute water accepting layer (4), nitrocellulose filter (6) layer middle part is sprayed with an anti-prostate characteristic antigen PSA monoclonal antibody, constitute detection line (3), between suction sample film (1) layer and nitrocellulose filter (6) layer, accompany collaurum film (2) layer, contain the gold mark probe of useful another anti-PSA Monoclonal Antibody in the collaurum film, go up adding one deck at suction sample film (1) and divide blood film (7) promptly to constitute the whole blood test test paper.

2, examination prostate cancer according to claim 1 and hyperplasia of prostate test paper,, it is characterized in that having in the test paper detection line (3), detection line (3) develops the color or does not develop the color, as measuring free PSA and total PSA ratio sign in human whole blood or the serum.

3, examination prostate cancer according to claim 1 and 2 and hyperplasia of prostate test paper is characterized in that the ratio that detection line (3) data are set to free PS A and total PSA is 10%, as colour developing and the cut off value that does not develop the color.

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