JPS5941141B2 - Filtering material for urine and body fluids - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a filter material for immunologically measuring trace biological components in urine and body fluids.

BACKGROUND ART It is widely practiced to measure trace amounts of hormones and other biological components secreted in urine and body fluids, and to use the results for clinical diagnosis or disease prognosis.

With recent advances in clinical chemistry and immunochemistry, methods for more precise measurement have been researched and developed, making a major contribution to the progress of clinical medicine. For example, by measuring human chorionic gonadotropin (hCG) in urine, confirmation of pregnancy, diagnosis of its condition, diagnosis of trophoblastic tumor, prognosis, etc. can be carried out, and by measuring human placental lactodiene (hPL), estriol, etc. Examples include diagnosing the condition of pregnancy, placenta, and fetus, and auxiliary diagnosis and prognosis of renal disease by measuring the amount of human fibrin body degradation product FDP in urine. Since trace measurements are required in these cases, methods that apply immunochemistry, such as direct agglutination reactions using blood cells (or latex) as carriers, or methods that use agglutination inhibition reactions, are generally used.

The former is hCG, FDP or human fibrinogen F.
The surface of red blood cells or latex particles is sensitized with an antibody obtained by immunizing animals such as rabbits, guinea pigs, and goats using G.g as an antigen, and this is mixed with test urine. It is known that the antigen corresponding to the antibody is present in the urine. In the latter method, blood cells or latex particles are sensitized with an antigen, and the corresponding antibody and test urine are mixed with the antigen.If the particles do not agglutinate, it is known that the antigen is present in the urine. . In both of these reactions, the minimum detection amount (sensitivity) can be adjusted by specifying the amount of antibody or antigen that sensitizes particles, so that not only qualitative detection but also quantitative detection is possible.
When even more sensitive and precise measurements are required, radioimmunoassay is used. This can be done, for example, by adding a certain amount of isotope-labeled antigen to the reaction system between the antigen in the test urine and its antibody, causing a competitive reaction, and measuring the amount of the labeled antigen bound to the antibody. It is used to know the amount of antigen. However, while these immune responses are highly specific, their reaction systems are susceptible to subtle influences.

In particular, urine contains complex components and has large individual differences, so when measuring trace components in urine immunologically, we are often faced with non-specific interference effects of urine components. or may be incorrectly judged as positive even though the FDP content is below the sensitivity. To avoid this, body fluids or urine are usually passed through or centrifuged before measurement. However, in the general detection method using paper, etc., adsorption loss of the analyte in urine to the paper occurs, and there is a risk of missing detection if the amount present is close to the detection sensitivity. Furthermore, when precise quantitative values were required, there was a risk of clinically significant misjudgment. It is also known that polyacrylonitrile synthetic fibers or cellulose acetate fibers can be used as filtering materials.

However, using these fibers, for example, 10 mIU/ml
It has been known that when a large amount of HCG is passed through, an adsorption loss of 10 to 22% occurs. The present inventors have previously discovered that the above-mentioned drawbacks of conventional filtering materials can be eliminated by treating the fibrous material with a specific substance (Japanese Patent Application Laid-Open No. 52-1979-1).
No. 83930). The present invention is a urine and body fluid filtration material for immunoassays consisting of cotton fibers treated with immunologically inert substances in the assay system. As a result of further research, the present inventors found that even when using synthetic resin foams or sintered bodies treated in the same way instead of cotton fibers, non-specific reactions contained in urine and body fluids were observed. It has been found that it is possible to advantageously remove substances that cause analyte and prevent adsorption loss of the substance to be measured.

The present invention is based on this knowledge, and consists of a synthetic resin foam or sintered body treated with an immunologically inert substance in a measurement system, and is a material for immunological measurement of urine and body fluids. It is.

The filter material of the present invention is produced by treating a synthetic resin foam or sintered body (hereinafter abbreviated as a synthetic resin porous body) with an immunologically inactive substance in a measurement system.

This treatment is preferably carried out by immersing the synthetic resin porous body in a solution of an inert substance (microadsorption), and then washing it with water or drying it without washing it with water. As the synthetic resin porous body, various types of synthetic resin substances having porosity are used.

Preferred are, for example, foams based on polyvinyl formal, sintered bodies of polyethylene, polypropylene, polyamide, and the like. It is particularly preferable that the synthetic resin porous body has continuous pores, high porosity, and low fluid resistance. This type of synthetic resin porous body is also suitable for handling large amounts of samples. The synthetic resin porous body can be manufactured by a known method, but commercially available products may also be used. Some immune reactions exhibit specific cross-reactions to the system.

For example, human luteinizing hormone HLH1 human follicle stimulating hormone (HFSH) for HCG, Fg for FDP, etc. cannot be used for each system. Furthermore, when measuring human biological components, serum proteins derived from humans are not preferred. Solutions containing metal ions that inhibit immune reactions are also not preferred. As the immunologically inactive substance in the measurement system, proteins such as serum albumin of animals such as cows, horses, guinea pigs, and rats are preferably used, and bovine serum albumin BSA is particularly preferred. Amino acids such as glycine and polypeptides are also used as inert substances. Furthermore, when a chelating agent such as ethylenediaminetetraacetic acid EDTA is used in combination with these inert substances, the effect of suppressing nonspecific reactions can be further improved. The inert substance is generally 0.05 to 1
%, especially 0.1 to 0.5% aqueous solution, and further add 1 to 20 wg/EDTA to this solution.
It is particularly preferable to add 2 to 10 mg/ml.

It is more preferable to use a buffer solution instead of an aqueous solution in order to take into account the auxiliary effect of an inert substance and to maintain a pH range suitable for immune reactions. The filtering material of the present invention may have any various shapes, such as blocks, cylinders, sheets, plates, etc.
It can be used in flakes, granules or other forms, and may be formed into these forms before or after treatment with an immunologically inert substance.

Since the urine and body fluids used in immunoassays are usually small quantities, a shape that is compatible with common screening equipment is preferred. For example, in the case of a glass road, you can use a sheet-like ▲ diaphragm like r paper, fit a conical diaphragm that matches the inner wall of the road into the road, or use a diaphragm of approximately the same diameter as the perforated plate. It is used by placing it on a circular sheet or disc. Alternatively, a cylindrical diaphragm may be closely inserted into the tubular portion of a load, dropper, or plastic tube. For example, a polyethylene tube with an inner diameter of about 5 Tm is filled with the present diaphragm to a length of 1 to 2 a1, and this tube is used by attaching it to the tip of a dropper. During normal use, the Honsawa filtration material swells slightly with urine and body fluids and comes into close contact with the load wall or tube wall. This material is used to remove human urine and body fluids such as cerebrospinal fluid, as well as to remove urine and body fluids from animals such as livestock, poultry, and pets.
May be used in excess.

When using the synthetic resin porous material according to the present invention, it is necessary to prevent adsorption loss of the analyte in urine and body fluids, and to prevent components (non-specific substances that cause reactions) can be removed.

This allows highly accurate quantitative results to be obtained, which can be used for clinical diagnosis. For example, in order to obtain data for diagnosing pregnancy, cancer, kidney diseases, and other endocrine diseases, trace amounts of biological components in urine and body fluids can be quantified with high precision. In the case of pregnancy diagnosis, it is performed by measuring a relatively large amount of HCG (11 V/ml or more), but in order to determine the prognosis of trophoblastic tumor, it is necessary to quantify a trace amount of HCG of about 501↓4d. By using the filtration material of the present invention, highly accurate results can be obtained even in such minute quantitative measurements. Example 1 A polyvinyl formal foam was prepared from BSAO. l% and E
DTA5ll! Glycine-NaOH containing g/ml
After soaking overnight in a buffer solution (pH 9.2, μ=0.24), the foam is then taken out and dried to obtain the ▲ filter material of the present invention.

Example 2 A polypropylene sintered body was prepared using BSAOO. After being immersed in a 5% aqueous solution overnight, it was dried to form a ▲filter material.

Example 3 A polyethylene sintered body is immersed overnight in a glycine buffer containing 0.5% rat serum albumin and then dried to obtain a filter material.

In the following test example, the sample was subjected to r-filtration as follows.

The untreated synthetic resin porous body and the ▲ filter material of the present invention obtained by processing in the same manner as in Examples 1 to 3 were treated in 1 to 1, respectively.
Fill a polyethylene tube with a length of 2C01 and an inner diameter of 5wi, and fit this tube into the tip of a dropper. P
After sucking the sample into the dropper through the filter material, remove the tube.] Vine.

The filtered sample in the syringe is then used for testing (measurement). In addition, in all test examples, a sample of 1.0 ml was used for each example. Test Example 1 Fifty healthy non-pregnant women's urine was tested by latex direct agglutination reaction (sensitivity: 1.01 V/ml).

As a result, 38 cases of excessive urination were found to be false positive or +
(Positive) (that is, a non-specific agglutination reaction not caused by HCG was shown). In addition, when urine was filtered through untreated polyvinyformal foam, polypropylene sintered body, or polyethylene sintered body, 3 cases, 5 cases, and 4 cases showed average to +, respectively. On the other hand, the urine filtered using the filtering material of the present invention obtained by treating the synthetic resin porous body in the same manner as in Examples 1 to 3 was found to be negative in all cases. )
showed that. These results show that non-specific reactions will occur if the urine is not thoroughly filtered, and that even if various synthetic resin porous materials are used, non-specific reactions will occur if the measurement system is not treated with an immunologically inactive substance. It is clear that this reaction cannot be completely eliminated. Test Example 2 Polyvinyl formal foam, polypropylene sintered body, or polyethylene sintered body was treated in the same manner as in Examples 1 to 3 for 100 urine samples from 5th week to 3rd month of normal pregnancy. Using the filtration material of the present invention, the obtained excess urine was subjected to latex agglutination reaction (sensitivity: 1.01-4d)
and hemagglutination inhibition reaction (sensitivity 0.51↓4J).

As a result, the positive rate was 97% for the polyethylene sintered body, 98% for the polypropylene sintered body, and 100% for the polyvinyl formal foam. Test Example 3 (HCGO by latex agglutination reaction) Adsorption test on P filter material) HCG was dissolved in the urine of healthy non-pregnant women at the concentrations shown in Table 1, and various synthetic resin porous bodies shown in Table 1 were used. ▲ and latex direct agglutination reaction (sensitivity 1.01
↓Tested according to 4d).

The results (changes in aggregation image) are shown in Table 1. The symbol 1 in the table means positive, 2 means false positive, and 1 means negative. As is clear from this result, the untreated synthetic resin porous material does not exhibit the specified sensitivity, and the use of the ▲filter material treated with an immunologically inactive substance in the measurement system according to the present invention For the first time, it showed the specified sensitivity of 1.01↓4d.
Test Example 4 (Adsorption test of HCGOP supermaterial using radioimmunoassay) HC was added to the urine of 5 healthy men at a concentration of 10 mIV/ml.
G was dissolved and passed through using the r-filtration material shown in Table 2 in the same manner as in Test Example 3, and the amount of HCG remaining in each urine was quantified by radioimmunoassay.

The results are shown in Table 2. The values in the table are the residual HCG charge (
%). From this result, it is known that the r-filtration material of the present invention exhibits an excellent improvement effect even in the case of a low concentration HCG solution.

Test Example 5 FDP in urine was tested by direct agglutination reaction using anti-human fibrinogen rabbit γ-globulin sensitized latex reagent (sensitivity 0.5 μg/ml).

As in Test Example 1, 40 out of 50 healthy men's urine showed non-specific agglutination images despite the amount of FDP present below the sensitivity in untreated hyperuria. On the other hand, urine filtered using the ▲filtration material of the present invention showed negative results in all cases with the amount of cellulose present below the sensitivity. From this, it is clear that even when quantifying the amount of FDP in urine, the urine must be filtered in advance.
Test example 6 (Adsorption test of FDP to ▲ supermaterial) FD was applied to five healthy male urine samples at a concentration of 100 μ [4d].
P was dissolved and filtered using the ▲ filter shown in Table 3, and the amount of FDP remaining in each filter was quantified by radioimmunoassay.

The results are shown in Table 3. The numbers in the table are the remaining FDP amount (
%). These results show that although FDP has greater adsorption than HCG, excellent improvement effects can be obtained by using the filtering material of the present invention.

Claims (1)

[Claims] 1. A filter material for urine and body fluids for immunological measurements, consisting of a synthetic resin foam or sintered body treated with an immunologically inert substance in the measurement system.