CA1141291A - Radioisotope muscle type aldolase antibody - Google Patents
Radioisotope muscle type aldolase antibodyInfo
- Publication number
- CA1141291A CA1141291A CA000372438A CA372438A CA1141291A CA 1141291 A CA1141291 A CA 1141291A CA 000372438 A CA000372438 A CA 000372438A CA 372438 A CA372438 A CA 372438A CA 1141291 A CA1141291 A CA 1141291A
- Authority
- CA
- Canada
- Prior art keywords
- muscle type
- type aldolase
- antibody
- human
- radioisotope
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/40—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against enzymes
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/573—Immunoassay; Biospecific binding assay; Materials therefor for enzymes or isoenzymes
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- Urology & Nephrology (AREA)
- General Health & Medical Sciences (AREA)
- Hematology (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- Organic Chemistry (AREA)
- Biotechnology (AREA)
- Physics & Mathematics (AREA)
- Cell Biology (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Biophysics (AREA)
- Food Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Analytical Chemistry (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
- Peptides Or Proteins (AREA)
Abstract
ABSTRACT OF THE DISCLOSURE:
Novel reagents and process for the correct and simple determination of human-muscle type aldolase. The reagent comprises radioisotope-labeled muscle type aldolase anti-body. The process comprises the following steps:
(a) contacting a specimen with insolubilized muscle type aldolase, followed by separation of the solid phase;
(b) contacting the solid phase with radioisotope-labeled muscle type aldolase antibody, followed by separa-tion of the solid phase; and (c) determining the radioactivity of the solid phase.
Novel reagents and process for the correct and simple determination of human-muscle type aldolase. The reagent comprises radioisotope-labeled muscle type aldolase anti-body. The process comprises the following steps:
(a) contacting a specimen with insolubilized muscle type aldolase, followed by separation of the solid phase;
(b) contacting the solid phase with radioisotope-labeled muscle type aldolase antibody, followed by separa-tion of the solid phase; and (c) determining the radioactivity of the solid phase.
Description
~14125'11 REAGENT AND METHOD FOR THE DETERMINATION
OF THE HUMAN-MUSCLE TYPE ALDOLASE
This invention relates to reagents and a method for the determination of the human-muscle type aldolase.
Aldolase is a general term of the enzymes which catalyze aldol condensation and cleavage between dihydro-xyacetone phosphate and a variety of aldehydes.
Aldolase according to this invention is meant for a fructose diphosphate aldolase (E.C. 4. 1. 2. 13. fructose-l, 6-phosphate D-glycelaldehyde-3-phosphate lyase) which reversibly decomposes fructose-1,6-diphosphate into dihydro-xyacetone phosphate and D-glycelaldehyde-3-phosphate. This enzyme occupies the main course of glycolysis system, and particularly, distrib~ites mainly in muscle, thereby contrib-uting importantly to the energy metabolism.
Mammal aldolase comprises three types of isoenzymes, that is, the muscle type (A type), the liver type (B type) and the brain type (C type). It is known that the construc-tion ratio of these isoenzymes varies in accordance with the metamorphosis of the living body, such as differentiation of the fetal liver in rat, canceration of organism in human, rat, and the like.
There have been conventionally known two methods for determining the human-muscle type aldolase, one of which is the electrophoretic method, and the other is a method which comprises measuring the decomposition activities of two substrates, that is, fractose-1,6-diphosphate (FDP) and fructose-l-phosphate (FIP) for aldolase, [FDP-ALD activity , ~
and FIP-ALD activity], thereby determining the activity ratio (FDP/FIP).
These methods, however, involve various problems.
In the electrophoretic method, the problem is that the migrated points of the human aldolase isoenzymes are so closely positioned one another that the discrimination is difficult and the precise quantitative determination can not be achieved.
In the measuring method of the activity ratio FDP/FIP, it is pointed out that the operation is particularly compli-cated for the measurement of the FIP-ALD activity, accompa-nied by no quantitatiVe determination.
Recently, there has been reported a method for determin-ing aldolase by means of radioimmunoassay through competition (double antibody method). Comparing with the aforementioned methods, this method is more excellent in the sensitivity and also in the quantification. However, this method also has some disadvantage that there is required a large amount of the second antibody for the separation of B and F, as well as handling procedure itself of the separation lS very complicated, resulting in the requ~ements of a long period of time and much labor cost. There is further observed an appearance of non-specific reaction.
The present invention relates to a radioisotope-labeled muscle type aldolase antibody and a method for the determi-nation of the human-muscle type aldolase by means of radio-immunoassay through so-called "Sandwich method" using the above-mentioned aldolase antibody as a reagent. According to this method, various defects in the methods of prior art can be dissolved or diminished. More particularly, the method of the present invention is remarkable in both the sensitivity for the determination an~d the quantitative matter. Also in the comparison with a double-antibody method, the method of the present invention is characterized in the points that the second antibody is not required; the separation procedure for B and F is simple and convenient;
and non-specific reaction is outstandingly low.
Accordins to so-called "Sandwich method" of the present invention, determination of human-muscle type aldolase is carried out through the following steps:
(a) contacting a specimen with an insolubilized muscle type aldolase antibody, followed by separation of the solid phase;
(b) contacting the solid phase with a radioisotope-labeled muscle type aldolase antibody, followed by separation of the solid phase; and (c~ determining the radioactivity of the solid phase.
The following will illustrate more particularly this method.
In the step (a), an antibody insolubilized by combina-tion with a carrier is incubated together with a specimen, such as serum, etc., so that the antigen (human muscle type aldolase) in the specimen may react with said insolubilized antibody. After the reaction is over, the reaction solution is removed and the solid phase is washed with buffer solu-tion, distilled water,physiological saline water or the like.
In the step (b), the solid phase obtained in the step (a) is incubated together with a radioisotope-labeled antibody, resulting in that said radioiso-tope-labeled anti-body combines with the antigen previously combined with the insolubilized antibody. After the reaction is over, the reaction solution is removed, and the solid phase is washed with a buffer solution, distilled water, physiological saline water or the like.
In the step (c), the radioactivity of the solid phase obtained in the step (b) is determined using a well-type scintilation counter, etc.
The radioactivity of the standard antigen is previously measured with said determination system, so as to set up a standard curve of the amount of antigen versus the radio-activity. The radioactivity of a specimen is measured with the same determination system, and then the amount of antigen in the specimen is determined from said standard curve.
Fig. 1 in the attached drawing shows a standard curve of the human-muscle type aldolase obtained by the radio-immunoassay in the following Exampie 2.
The following description illustrates the reagent for determination to be used with the present method and the muscle type aldolase antibody to be used with the production of the reagent.
A) The muscle type aldolase antibody Antiserum is obtained by immunizing the muscle type aldolase of a mammal such as human, rabbit, dog, monkey, etc. to the other animal.
The immune animal is preferably selected from the species of birds, because the muscle type aldolases do not ~141~
show significant immunological distinction between the mammals. Illustrative of the preferred birds include, for example, fowl, turkey, duck and the like.
In the purification of the resulting antiserum, there can be used a method which comprises adding the inactivated serum of the animal which is the source of the muscle type aldolase, and removing by absorption the other impure anti-body; an affinity chromatography using support combined the muscle type aldolase; and the like.
B) Radioisotope-labeled muscle type aldolase antibody The labeling of muscle type aldolase antibody by means of radioisotope can be achieved by any conventional methods.
For example, the labeling is carried out by chloramine-T method using 125I, 13~I, etc. as the radioisotope, and the like method.
C) The insolubilized muscle type aldolase antibody As a carrier, there are used an insoluble solid which is combinative with an antibody. Illustrative of such solid include, for example, polystyrene, cellulose, agarose, glass, bridged dextran, metal, etc. There may be mentioned a form such as tube, microtiter plate, powder, sphere, disc, plate, foil, etc.
In the case of the polystyrene microtiter plate, the antibody may combine with the wall surface of the plate, when the antibody is properly diluted with a buffer solution or the like, the diluted solution is then added to the plate, and the whole is finally allowed to stand.
The above descriptions have been made to the sandwich method, but the human muscle type aldolase can be also determined by the immunoradiometric assay using radioisotope-labeled muscle type aldolase antibody which is the reagent of the present invention. This method comprises reacting the radioisotope-labeled antibody with a specimen (antigen), followed by separation of the radioisotope-labeled antibody-antigen reactant (B) from unreacted radioisotope-labeled anti-body (F), and determining the radioactivity of either said (B) or (F). As the method for the separation of (B) and (F), there may be mentioned the gel filtration method, the absorp-tion method by means of insolubilized antigen, etc.
Following experiments and examples will further illus-trate this invention.
Experiment 1 Human muscle type aldolase antibody The human-muscle type aldolase (1 mg) was dissolved in physiological salt water (0.5 ml), and to the solution was admixed the equal volume of Freund complete adjuvant. The product was immunized by subcutaneous injection to a fowl (~hite Leghorn) at the intervals of one week, so that the injections may amount to five times. One week after the last immunization, the blood was collected from the fowl to obtain the anti-serum.
Sepharose 4BTM (Pharmacia Fine Chemicals AB; Agarose) (10 g) which was activated with bromo-cyanide, and the human-muscle type aldolase (10 mg) were allowed to stand at 4C overnight in 15 ml of 0.1M sodium carbonate buffer solu-tion (pH 9.0).
The resulting Sepharose 4B gel combined with the human-muscle type aldolase was packed into a column, and washed with 0.05 M tris-hydrochloric acid buffer solution (pH 8.0) containing 0.5 N sodium chloride. To this column was added the above-mentioned human-muscle type aldolase anti-serum, and the anti-serum was flowed out with 0.05 M tris-hydro-chloric acid buffer solution (pH 8.0) containing 0.5 N
sodium chloride. Thereafter, 0.1 M aqueous sodium carbonate solution was added to the column to elute the human-muscle type aldolase antibody. The fraction of antibody eluate was subjected to dialysis with 0.05 M tris-hydrochloric acid buffer solution (pH 8.0) to obtain 3 mg of the human-muscle type aldolase antibody.
Experiment 2 _nsolubilized human-muscle type aldolase antibody Into each of the holes provided on a polystylene micro-titer plate, was added every 200 ~1 of solution of the human-muscle type aldolase antibody which was adjusted to OD280 m~
=0.10, and the whole was then allowed to stand overnight at 4C. The solution was removed from the plate. The plate was washed with distilled water to obtain the microtiter plate combined with the human-muscle type aldolase antibody.
5I-labeled human muscle type aldolase antibody Na I of 250 ~ curie (10 ~1), 0.5 M phosphate buffer (pH 7.4)(12.5 ~1), human muscle type aldolase antibody (1.85 mg/ml)(10 ~1) and Chloramine-T (3 mg/ml)(12.5 ~1) were reacted one another for 20 seconds in a small test tube coated with silicone. After the reaction was over, sodium metabisulfite (6 mg/ml)(25 ~1) were added to the mixture, so as to cease the reaction. To the resulting mixture were added potassium iodide (50 mg/ml)(12.5 ~1) and 2~ egg albumin (250 ~1) in 0.05 M phosphate buffer. The whole was then subjected to gel-filtration by means of Sephadex G-25TM
(Pharmacia Fine Chemicals AB; bridged-dextran), so that the free 125I may be removed. There was thus obtained l25I-labeled human muscle type aldolase antibody, the specific activity thereof being 11.6 ~ curie/~g.
Radioimmunoassay (Sandwich method) Into each of the holes which were provided on microtiter plate combined with human muscle type aldolase antibody, 100 ~ of the standard solution of the human muscle type aldolase were respectively added. The whole was reacted at 37C for
OF THE HUMAN-MUSCLE TYPE ALDOLASE
This invention relates to reagents and a method for the determination of the human-muscle type aldolase.
Aldolase is a general term of the enzymes which catalyze aldol condensation and cleavage between dihydro-xyacetone phosphate and a variety of aldehydes.
Aldolase according to this invention is meant for a fructose diphosphate aldolase (E.C. 4. 1. 2. 13. fructose-l, 6-phosphate D-glycelaldehyde-3-phosphate lyase) which reversibly decomposes fructose-1,6-diphosphate into dihydro-xyacetone phosphate and D-glycelaldehyde-3-phosphate. This enzyme occupies the main course of glycolysis system, and particularly, distrib~ites mainly in muscle, thereby contrib-uting importantly to the energy metabolism.
Mammal aldolase comprises three types of isoenzymes, that is, the muscle type (A type), the liver type (B type) and the brain type (C type). It is known that the construc-tion ratio of these isoenzymes varies in accordance with the metamorphosis of the living body, such as differentiation of the fetal liver in rat, canceration of organism in human, rat, and the like.
There have been conventionally known two methods for determining the human-muscle type aldolase, one of which is the electrophoretic method, and the other is a method which comprises measuring the decomposition activities of two substrates, that is, fractose-1,6-diphosphate (FDP) and fructose-l-phosphate (FIP) for aldolase, [FDP-ALD activity , ~
and FIP-ALD activity], thereby determining the activity ratio (FDP/FIP).
These methods, however, involve various problems.
In the electrophoretic method, the problem is that the migrated points of the human aldolase isoenzymes are so closely positioned one another that the discrimination is difficult and the precise quantitative determination can not be achieved.
In the measuring method of the activity ratio FDP/FIP, it is pointed out that the operation is particularly compli-cated for the measurement of the FIP-ALD activity, accompa-nied by no quantitatiVe determination.
Recently, there has been reported a method for determin-ing aldolase by means of radioimmunoassay through competition (double antibody method). Comparing with the aforementioned methods, this method is more excellent in the sensitivity and also in the quantification. However, this method also has some disadvantage that there is required a large amount of the second antibody for the separation of B and F, as well as handling procedure itself of the separation lS very complicated, resulting in the requ~ements of a long period of time and much labor cost. There is further observed an appearance of non-specific reaction.
The present invention relates to a radioisotope-labeled muscle type aldolase antibody and a method for the determi-nation of the human-muscle type aldolase by means of radio-immunoassay through so-called "Sandwich method" using the above-mentioned aldolase antibody as a reagent. According to this method, various defects in the methods of prior art can be dissolved or diminished. More particularly, the method of the present invention is remarkable in both the sensitivity for the determination an~d the quantitative matter. Also in the comparison with a double-antibody method, the method of the present invention is characterized in the points that the second antibody is not required; the separation procedure for B and F is simple and convenient;
and non-specific reaction is outstandingly low.
Accordins to so-called "Sandwich method" of the present invention, determination of human-muscle type aldolase is carried out through the following steps:
(a) contacting a specimen with an insolubilized muscle type aldolase antibody, followed by separation of the solid phase;
(b) contacting the solid phase with a radioisotope-labeled muscle type aldolase antibody, followed by separation of the solid phase; and (c~ determining the radioactivity of the solid phase.
The following will illustrate more particularly this method.
In the step (a), an antibody insolubilized by combina-tion with a carrier is incubated together with a specimen, such as serum, etc., so that the antigen (human muscle type aldolase) in the specimen may react with said insolubilized antibody. After the reaction is over, the reaction solution is removed and the solid phase is washed with buffer solu-tion, distilled water,physiological saline water or the like.
In the step (b), the solid phase obtained in the step (a) is incubated together with a radioisotope-labeled antibody, resulting in that said radioiso-tope-labeled anti-body combines with the antigen previously combined with the insolubilized antibody. After the reaction is over, the reaction solution is removed, and the solid phase is washed with a buffer solution, distilled water, physiological saline water or the like.
In the step (c), the radioactivity of the solid phase obtained in the step (b) is determined using a well-type scintilation counter, etc.
The radioactivity of the standard antigen is previously measured with said determination system, so as to set up a standard curve of the amount of antigen versus the radio-activity. The radioactivity of a specimen is measured with the same determination system, and then the amount of antigen in the specimen is determined from said standard curve.
Fig. 1 in the attached drawing shows a standard curve of the human-muscle type aldolase obtained by the radio-immunoassay in the following Exampie 2.
The following description illustrates the reagent for determination to be used with the present method and the muscle type aldolase antibody to be used with the production of the reagent.
A) The muscle type aldolase antibody Antiserum is obtained by immunizing the muscle type aldolase of a mammal such as human, rabbit, dog, monkey, etc. to the other animal.
The immune animal is preferably selected from the species of birds, because the muscle type aldolases do not ~141~
show significant immunological distinction between the mammals. Illustrative of the preferred birds include, for example, fowl, turkey, duck and the like.
In the purification of the resulting antiserum, there can be used a method which comprises adding the inactivated serum of the animal which is the source of the muscle type aldolase, and removing by absorption the other impure anti-body; an affinity chromatography using support combined the muscle type aldolase; and the like.
B) Radioisotope-labeled muscle type aldolase antibody The labeling of muscle type aldolase antibody by means of radioisotope can be achieved by any conventional methods.
For example, the labeling is carried out by chloramine-T method using 125I, 13~I, etc. as the radioisotope, and the like method.
C) The insolubilized muscle type aldolase antibody As a carrier, there are used an insoluble solid which is combinative with an antibody. Illustrative of such solid include, for example, polystyrene, cellulose, agarose, glass, bridged dextran, metal, etc. There may be mentioned a form such as tube, microtiter plate, powder, sphere, disc, plate, foil, etc.
In the case of the polystyrene microtiter plate, the antibody may combine with the wall surface of the plate, when the antibody is properly diluted with a buffer solution or the like, the diluted solution is then added to the plate, and the whole is finally allowed to stand.
The above descriptions have been made to the sandwich method, but the human muscle type aldolase can be also determined by the immunoradiometric assay using radioisotope-labeled muscle type aldolase antibody which is the reagent of the present invention. This method comprises reacting the radioisotope-labeled antibody with a specimen (antigen), followed by separation of the radioisotope-labeled antibody-antigen reactant (B) from unreacted radioisotope-labeled anti-body (F), and determining the radioactivity of either said (B) or (F). As the method for the separation of (B) and (F), there may be mentioned the gel filtration method, the absorp-tion method by means of insolubilized antigen, etc.
Following experiments and examples will further illus-trate this invention.
Experiment 1 Human muscle type aldolase antibody The human-muscle type aldolase (1 mg) was dissolved in physiological salt water (0.5 ml), and to the solution was admixed the equal volume of Freund complete adjuvant. The product was immunized by subcutaneous injection to a fowl (~hite Leghorn) at the intervals of one week, so that the injections may amount to five times. One week after the last immunization, the blood was collected from the fowl to obtain the anti-serum.
Sepharose 4BTM (Pharmacia Fine Chemicals AB; Agarose) (10 g) which was activated with bromo-cyanide, and the human-muscle type aldolase (10 mg) were allowed to stand at 4C overnight in 15 ml of 0.1M sodium carbonate buffer solu-tion (pH 9.0).
The resulting Sepharose 4B gel combined with the human-muscle type aldolase was packed into a column, and washed with 0.05 M tris-hydrochloric acid buffer solution (pH 8.0) containing 0.5 N sodium chloride. To this column was added the above-mentioned human-muscle type aldolase anti-serum, and the anti-serum was flowed out with 0.05 M tris-hydro-chloric acid buffer solution (pH 8.0) containing 0.5 N
sodium chloride. Thereafter, 0.1 M aqueous sodium carbonate solution was added to the column to elute the human-muscle type aldolase antibody. The fraction of antibody eluate was subjected to dialysis with 0.05 M tris-hydrochloric acid buffer solution (pH 8.0) to obtain 3 mg of the human-muscle type aldolase antibody.
Experiment 2 _nsolubilized human-muscle type aldolase antibody Into each of the holes provided on a polystylene micro-titer plate, was added every 200 ~1 of solution of the human-muscle type aldolase antibody which was adjusted to OD280 m~
=0.10, and the whole was then allowed to stand overnight at 4C. The solution was removed from the plate. The plate was washed with distilled water to obtain the microtiter plate combined with the human-muscle type aldolase antibody.
5I-labeled human muscle type aldolase antibody Na I of 250 ~ curie (10 ~1), 0.5 M phosphate buffer (pH 7.4)(12.5 ~1), human muscle type aldolase antibody (1.85 mg/ml)(10 ~1) and Chloramine-T (3 mg/ml)(12.5 ~1) were reacted one another for 20 seconds in a small test tube coated with silicone. After the reaction was over, sodium metabisulfite (6 mg/ml)(25 ~1) were added to the mixture, so as to cease the reaction. To the resulting mixture were added potassium iodide (50 mg/ml)(12.5 ~1) and 2~ egg albumin (250 ~1) in 0.05 M phosphate buffer. The whole was then subjected to gel-filtration by means of Sephadex G-25TM
(Pharmacia Fine Chemicals AB; bridged-dextran), so that the free 125I may be removed. There was thus obtained l25I-labeled human muscle type aldolase antibody, the specific activity thereof being 11.6 ~ curie/~g.
Radioimmunoassay (Sandwich method) Into each of the holes which were provided on microtiter plate combined with human muscle type aldolase antibody, 100 ~ of the standard solution of the human muscle type aldolase were respectively added. The whole was reacted at 37C for
2 hours. After the reaction was over, the supernatant layer were removed, and the remainder was washed thrice with physiological saline solution. To the washed holes was then respectively added 100 ~1 of 125I-labeled human muscle type aldolase antibody (about 200,000 cpm). The whole was reacted at room temperature for 16 hours. After the reaction was over, the supernatant layer was removed. The remainder was washed thrice with physiological saline solution.
After the contained water was thoroughly exhausted, the plate was cut, and the respective hole portions were sepa-rated one another. The individual aliquots were respective-ly placed in small test tubes. Radioactivity was counted in a well-type scintilation counter.
As shown in Figure 1, there was obtained the standard curve wherein about 8 - 1000 ng/ml were plotted as measur-able ranges. In the Figure, the horizontal axis (logarithm) indicates the concentration (ng/ml) of the human muscle type aldolase, and the vertical axis indicates the radioactivity (cpm).
~ luman muscle type aldolases in various human serums were then determined through the above assay system. There were thus observed the data that the human muscle type aldolase of 25 normal subject amount to 165 + 40 ng/ml, all of which were less than 210 ng/ml. On the contrary, the human muscle type aldolase of 20 cancer patients showed higher values than 210 ng/ml, excludir.g two examples. The 20 cancer patients comprise 7 hepatomae, 8 gastric cancers,
After the contained water was thoroughly exhausted, the plate was cut, and the respective hole portions were sepa-rated one another. The individual aliquots were respective-ly placed in small test tubes. Radioactivity was counted in a well-type scintilation counter.
As shown in Figure 1, there was obtained the standard curve wherein about 8 - 1000 ng/ml were plotted as measur-able ranges. In the Figure, the horizontal axis (logarithm) indicates the concentration (ng/ml) of the human muscle type aldolase, and the vertical axis indicates the radioactivity (cpm).
~ luman muscle type aldolases in various human serums were then determined through the above assay system. There were thus observed the data that the human muscle type aldolase of 25 normal subject amount to 165 + 40 ng/ml, all of which were less than 210 ng/ml. On the contrary, the human muscle type aldolase of 20 cancer patients showed higher values than 210 ng/ml, excludir.g two examples. The 20 cancer patients comprise 7 hepatomae, 8 gastric cancers,
3 large intestine cancers, and 2 pancreatic cancers.
Claims (4)
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:
1. A reagent for the determination of human-muscle type aldolase which comprises radioisotope-labeled muscle type aldolase antibody.
2. The reagent according to claim 1, wherein said radioisotope is 125I.
3. The reagent according to claim 1, wherein said muscle type aldolase antibody is human-muscle type aldolase antibody.
4. A process for the determination of human-muscle type aldolase which comprises the following steps:
(a) contacting a specimen with an insolubilized muscle type aldolase antibody, followed by separation of the solid phase;
(b) contacting the solid phase with radioisotope-labeled muscle type aldolase antibody, followed by separa-tion of the solid phase; and (c) determining the radioactivity of the solid phase.
(a) contacting a specimen with an insolubilized muscle type aldolase antibody, followed by separation of the solid phase;
(b) contacting the solid phase with radioisotope-labeled muscle type aldolase antibody, followed by separa-tion of the solid phase; and (c) determining the radioactivity of the solid phase.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP27,996/80 | 1980-03-07 | ||
| JP2799680A JPS56125667A (en) | 1980-03-07 | 1980-03-07 | Reagent for measurement and measuring method for human muscle type aldolase |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1141291A true CA1141291A (en) | 1983-02-15 |
Family
ID=12236425
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000372438A Expired CA1141291A (en) | 1980-03-07 | 1981-03-06 | Radioisotope muscle type aldolase antibody |
Country Status (10)
| Country | Link |
|---|---|
| JP (1) | JPS56125667A (en) |
| BE (1) | BE887828A (en) |
| CA (1) | CA1141291A (en) |
| CH (1) | CH651933A5 (en) |
| DE (1) | DE3107268A1 (en) |
| FR (1) | FR2477573A1 (en) |
| GB (1) | GB2071318B (en) |
| IT (1) | IT1136853B (en) |
| NL (1) | NL8101068A (en) |
| SE (1) | SE8101404L (en) |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4649039A (en) * | 1984-07-03 | 1987-03-10 | E. I. Du Pont De Nemours And Company | Radiolabeling of methionine-containing proteins and peptides |
| JPH0672893B2 (en) * | 1985-08-16 | 1994-09-14 | 富士薬品工業株式会社 | Determination of human prolyl hydroxylase in human blood by radioimmunoassay. |
| WO2006067792A2 (en) * | 2004-12-22 | 2006-06-29 | Yeda Research And Development Co. Ltd. At The Weizmann Institute Of Science | Aldolase autoantigens useful in diagnosis and treatment of alzheimer's disease |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5648894A (en) * | 1979-09-25 | 1981-05-02 | Eisai Co Ltd | Reagent for measuring human muscular aldolase and its determination |
-
1980
- 1980-03-07 JP JP2799680A patent/JPS56125667A/en active Pending
-
1981
- 1981-02-26 DE DE19813107268 patent/DE3107268A1/en not_active Withdrawn
- 1981-03-02 CH CH1389/81A patent/CH651933A5/en not_active IP Right Cessation
- 1981-03-03 FR FR8104174A patent/FR2477573A1/en active Pending
- 1981-03-04 IT IT20117/81A patent/IT1136853B/en active
- 1981-03-04 SE SE8101404A patent/SE8101404L/en unknown
- 1981-03-05 NL NL8101068A patent/NL8101068A/en not_active Application Discontinuation
- 1981-03-06 GB GB8107059A patent/GB2071318B/en not_active Expired
- 1981-03-06 CA CA000372438A patent/CA1141291A/en not_active Expired
- 1981-03-06 BE BE0/204036A patent/BE887828A/en not_active IP Right Cessation
Also Published As
| Publication number | Publication date |
|---|---|
| IT8120117A0 (en) | 1981-03-04 |
| CH651933A5 (en) | 1985-10-15 |
| NL8101068A (en) | 1981-10-01 |
| BE887828A (en) | 1981-07-01 |
| SE8101404L (en) | 1981-09-08 |
| GB2071318B (en) | 1983-08-24 |
| FR2477573A1 (en) | 1981-09-11 |
| IT1136853B (en) | 1986-09-03 |
| JPS56125667A (en) | 1981-10-02 |
| GB2071318A (en) | 1981-09-16 |
| DE3107268A1 (en) | 1982-01-07 |
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