JPS6027375B2 - Device for the preparation of measuring fluids for use in optical analyzers - Google Patents

Description

[Detailed description of the invention]

The present invention relates to a device for the preparation of a measurement fluid for use in an optical analysis device, including a closed container filled with a precisely prepared reaction solution and a sample fluid introduced into the container for dispensing the reaction solution. It is based on capillary tubes. The invention can be particularly applied to analytical testing of bodily fluids. “Readytests” are increasingly being offered in the fields of hematology, clinical chemistry and biochemistry research, and these “readytests” provide users with hematological information without even the need to add reagents. , allowing clinical chemical and biochemical parameters to be measured quickly and easily. These “instant tests” are also “mono tests”, “
Also known as "single test" or "single glass test", it is characterized in that the labile components of the reagent for immediate use can be stored, usually in lyophilized form, in glass or plastic containers. Before the reagent is ready for use, a solvent is added which may contain further additional substances required for the complete reagent. The reagents to be provided may be present in glass or plastic containers.The sample to be analyzed, e.g. fluid, blood, blood count or serum, is prepared in a manner that ensures accurate formulation, usually by pipette. The reagent-sample mixture is then placed in a measurement container, usually glass or quartz, and analyzed with a measurement device, such as a photometer.Closeable containers have recently been identified and 242226), which is designed as a plastic disposable measuring container (preferably made of polystyrene) and is hermetically closed by a plastic cover. The cover is bonded to the container by gluing, for example by ultrasound. The cover has a predetermined break point that can be broken by a pin. The sample to be analyzed is thus made on the cover of the container. can be introduced into the container through a closed closure. The container itself already contains the complete reagent fluid.
For photometric measurements of ber), for example, the user only needs to introduce five volumes of blood into the container through an opening in the cover using a volumetrically calibrated glass capillary tube. Photometric measurements can occur as soon as the blood is thoroughly mixed with the reagents.
This method is particularly good because it does not require the user to add or dissolve reagents or require auxiliary tools for analysis such as pipettes. This considerably simplifies the analysis so that the method can be performed accurately even by untrained personnel. The hitherto known methods mentioned above have the following drawbacks: During the "immediate test" it is usually still necessary to carry out movements with a pipette. For this purpose, expensive and sensitive auxiliary equipment such as Eppendorf-Pipette
et) is required. The use of pipettes itself represents a source of error. The required solvents, e.g. distilled water, are not always available in the required quantities (e.g. quartz-
Distilled water). The complete reagent or complete reagent-sample mixture must be transferred to a measurement container, for example a glass container, before the actual measurement can be carried out. This is an additional operating step that introduces additional possibilities for error, such as containers that are not completely clean or not completely dried (resulting in volume errors) or residues or detergent in the containers. It is. Overall, these "instant tests" are too complex to be called "instant tests," and there are many sources of error that make these tests special. It cannot be performed by someone without proper training. The test described in German Patent Application No. 242-66 substantially meets the requirements of an "immediate test". The user does not have to dissolve reagents or precisely mix various solutions or transfer a prepared reagent-sample mixture to a measurement vessel. The user simply needs to introduce an accurately weighed sample to be analyzed into the container, which can be easily done, for example, by a volumetric capillary tube, and then the container is transferred to the photometer. can be done. Possible sources of error that may occur in such operations are thus eliminated from the beginning, and the analysis process is simplified so that even untrained persons can carry out the analysis accurately. Unfortunately, however, only a few reagents for biochemical, hematological and clinical chemistry analyzes remain usable for long periods of time (6 months). This means that
This is particularly true of modern enzymatic methods, such as glucose-oxidase/peroxidase, hexokinase/glucose-6-phosphate dehydrogenase or glucose-oxidase/peroxidase,
This applies to methods for measuring dish sugar levels using dehydrogenase-mutaratactic enzymes, for enzymatic determination of uric acid, urea, triglycerides and cholesterol, and of course for determining the activity of the enzyme itself. However, non-enzymatic assay reagents remain unused for long periods of time (
(6 months) will not remain usable. This is true, for example, of Jendrassik and G.
Determination of bilirubin using diazotized sulfanilic acid according to [2] or using 2,5-dichlorophenyludiazonium salt according to Wahle [3] and picric acid/NaOH similarly according to Popper [4]
This applies to the measurement of creatinine using The use of disposable measuring containers is even more problematic. Glass containers are excluded as a material for disposable measurement containers because they are too expensive for the quality required for photometric measurements. The use of plastics, such as polystyrene, allows for inexpensive mass production of disposable measuring containers, but is still associated with problems regarding reagent durability. Polystyrene is not absolutely impermeable to water vapor and unless suitable early measures are found, the volume of reagent fluid may vary in the closed container during the storage period. Erroneous results will be obtained due to the fact that the analysis to be performed depends on the mixing ratio being precisely known. Polystyrene is also permeable to many gases. This is a disadvantage, for example, to the durability of reagents for determining hemoglobin as haemigobin cyanide. This reagent consists of a buffer substance and erythroditene (er×hro
In addition to detergents for hemolysis of hemoglobin, it contains potassium hexacyanoferrite(m), which oxidizes hemoglobin to hemiglobin, and KCN, which converts hemiculobin to hemiglobin cyanide. Hemig. Vincyanide is measured photometrically. KCN contained in reagent
is in equilibrium with HCN in the gas above the fluid. However, since HCN can diffuse through the polystyrene walls of the container, the CN- content of the reagent decreases with a half-life of about 4 weeks. Therefore, the reagents are not nearly durable enough to be marketed. Therefore, the object of the present invention is to
Developing an "instant test" which overcomes the above-mentioned shelf life problems and in which prepared plastic disposable containers can be used without any additional equipment such as a pipette and with special training by the user. The objective is to allow accurate analysis results to be obtained in a short period of time. According to the invention, for use in an optical analysis device comprising a precisely prepared closed container containing a reaction solution and a capillary tube insertable into the container for dispensing a sample fluid into the reaction solution. In the apparatus for the preparation of a measuring fluid, the optical properties of the measuring fluid are such that at least one of the reactants required for the formation of the measuring fluid from the reaction solution is
Use in the optical analysis device, characterized in that it is prepared by being placed as a solid phase in the capillary tube used for preparing the sample fluid and/or in a second capillary tube that can be introduced into the container. A device is provided for the preparation of a measurement fluid for. The reactants are preferably placed on the inner wall of the capillary tube in finely divided form. In this way, the range of reactants can be expanded considerably. Finally, not all reactants can be stored together in one solution, since they become unstable in a short period of time or react with each other in this combined form. This problem is neatly solved by storing such unstable components in capillaries. Reactions that were hitherto considered impenetrable to the analytical method described in DE 242 226-1 due to the nonlocalization of the reactants can be achieved in this way. Capillaries coated with one reactant and stored dry can remain stable for long periods of time (6 months or more), whereas reactants that are stable in solution can be precisely prepared and delivered to the measuring vessel. obtain. The technician may: a Take a sample (fluid, blood, blood, serum) through a lined capillary tube and introduce the sample-filled capillary tube into a plastic disposable container. The complete reagent is thus formed in very simple and quick operating steps to precisely prepare the sample to be analyzed. If desired, after a certain reaction time the sample can be analyzed with a measuring device (eg photometer). In this case, of course, the capillary used should be calibrated in volume and the components of the reagent applied to the inner surface of the capillary should not change the total volume. b Place the coated capillary tube into the prepared plastic disposable measuring container to make the complete reagent. The sample to be analyzed next (fluid, blood, blood bran, serum)
may be added (e.g. using a volumetric capillary tube)
. If desired, after a certain reaction period, the sample can be analyzed with a measuring device (eg, a photometer). If a control value for a reagent has to be determined during the test, this value can be obtained before addition of the sample to be analyzed. When a reference value for a sample is to be determined, it is preferred that the reference value is determined when the sample is first placed in a prepared plastic disposable measurement container and the complete reagent is added to the coated capillary tube. is formed and then the analytical value is measured. In this way both the sample control value and the analytical value are 1.
can be measured in plastic disposable measuring containers. This eliminates the use of extra containers and the amount of sample that would be required to measure sample control values. c. Using a capillary tube with a coated inner surface, take up another labile component, for example an enzyme suspension or a suspension of an enzyme mixture, and place it in a prepared plastic disposable measuring container to form the complete reagent. Addition of the sample can occur before or after adding the labile component depending on whether reagent control values or sample control values are to be measured. It has been found that the method shown in a) gives particularly good results when, for example, hemoglobin is determined as hemoglobin cyanide. Place the traces of reagent 1.2 completely isolated from KCN into a plastic disposable measuring container. KCN (75 French grams per capillary tube) was placed in a finely pulverized state on the inner surface of a capillary tube with a capacity scale of 5 mu. A sample, in this case blood, is taken through this capillary. The capillary is automatically filled by capillary action as soon as the capillary is brought into contact with blood. KCN readily dissolves in the blood and does not affect the measurement accuracy of sample dispersion. Once the capillary tube is filled with blood, the capillary tube is then introduced into a plastic disposable measuring container. The contents of the capillary tube are mixed with the contents of the container by gently shaking the measuring container, so that the reagents are completed at the same time. After 3 minutes of reaction time, the hemoglobin content of the sample can be measured directly with a photometer. The method described in b. is found to yield particularly good results when bilirubin is determined using 3,5-dichlorophenyldiazonium salt according to Wahleseld et al. [3]. During this measurement, the sample control value must be taken into account.
Since a relatively large amount of sample is required for the determination of pirirubin content (sample control and analysis volumes require 50 mu of each serum), the measurement of sample control and analysis values can be carried out in one measuring vessel. It is particularly advantageous to obtain a sample of 50 AI (i.e. with sample 50AI).
1.25 volumes of complete reagent apart from the 2,5-dichlorophenyldiazonium salt are delivered to a plastic disposable measuring container. A sample (for example, 50 ml of serum) is placed in a container using a capillary tube, and the container is gently shaken. Reagent control values can then be measured with a photometer. The capillary tube with the 2,5-dichlorophenyldiazonium salt placed on its inner surface is then introduced into the container and gently shaken. After the reaction time (1 female h) has elapsed, the analytical values are determined with a photometer. The result of the bilirubin content of the sample is obtained by subtracting the sample control value from the analytical value and multiplying the difference by a defined factor. The method described in c can be used in cases where some of the components required for the measurement are unstable in solution, e.g. in the Roschlau method with a consequent star-colored reaction according to Trinde et al. [6].
It is preferably used for enzymatic cholesterol measurement according to [5]. 1.5' phosphate buffer/phenolomethanol/
The hydroxy-polyethoxide dodecane mixture is placed in a sealed plastic disposable measuring container. 4-aminophenazone reacts with phenol and further reacts with KO2 generated during the enzymatic reaction of cholesterol to give a color component [4-(p-benzoquinone monoimino)-phenazone] that is measured by photometry. . 4-Aminophenazone is placed as a solid on the inner surface of a glass or plastic capillary. This is because 4-aminophenazone, even during storage, together with phenol in solution forms the colored component that is to be measured after the enzymatic reaction. A capillary tube coated with 4-aminophenazone can be used, for example, to introduce a suspended enzyme mixture (approximately 20 rl of cholesterol esterase/cholesterol oxidase/peracid Q enzyme) in ammonium sulfate into a plastic disposable measuring container. used. The contents of the capillary tube are mixed with the contents of the container by gentle shaking and the control value of the reagent is determined photometrically. The sample to be analyzed (fluid, serum, blood acid) is added via a volumetric capillary, the container is gently shaken, and after a reaction time of about 15 minutes the analytical value is determined with a photometer. The resulting cholesterol content of the sample is obtained by subtracting the reagent control value from the analytical value and multiplying the difference by a defined factor. The present invention described above is commercially available.
Hematological, clinical chemistry, and biochemistry tests that are in fact "instant tests," unlike the "mono-st", "singeglass test" or "single test". The use of the measurement container of the invention allows the same container to be used for sample preparation and measurement. Therefore, the user no longer has to mix the sample with reagents in a special reaction vessel and then transfer this to the measurement vessel. This results in cost savings (no reaction vessel is required) and simplification of operation. Since the measuring vessel contains all the liquid components of the reaction solution in precise quantities and the labile components are placed on the inner surface of the capillary tube (glass or plastic) in precise quantities, the reaction The solution can be easily and easily completed by inserting a capillary tube into a plastic disposable measuring container. Errors in measuring and dissolving unstable components are thus eliminated. The user does not have to carry out the steps with a pipette. The analysis is therefore easier, faster and more accurate. It is particularly advantageous to apply the labile component to the inner surface of the capillary. Small amounts (micrograms) can be placed precisely into the capillary. No excipients (such as binders and fillers required for tablet manufacturing) that would interfere with reactions that are currently sensitive to excipients are required. dissolves rapidly (approximately 1
tablets require, for example, 10 minutes for complete dissolution. Turbidity can now be caused by additives, thereby avoiding photometric analysis, which is more difficult. Direct contact of hazardous substances with the skin (fingers) is prevented. Even unstable components in the reaction solution can last for several years if stored in a dry, cool place, greatly increasing the durability of the test. Reaction time and therefore analysis time can be reduced. For example, when measuring hemoglobin by the hemimoglobin cyanide method, the pH value of the solution is adjusted to 7.2.
This is a compromise between the reaction rate, which decreases as the pH value of the solution increases, and the durability of the solution, which increases as the pH value increases. This is because HCN is driven out of solution more quickly at lower pH values, thus making the reaction solution quickly unusable. Reaction time: pH value 7.2
That will be 3 minutes. At a pH value of 6.8, the reaction time is only 1 minute, but
The solution is no longer stable for long periods of time. However, K
When a capillary tube coated inside with CN is used, the p of the solution
The H value can be adjusted to, for example, 6.8 and the stability is good, so that the reaction time can be significantly shortened. Since no auxiliary vehicles such as pipettes or metering cylinders are required to complete the reagents of these solution instant tests, these tests are ideally suited for mobile applications, such as rescue helicopters, ambulances or house calls. Portable battery-operated photometers (rMIOOO on Compurmini-phobme) are available for these tests. The use of plastic disposable measuring containers also offers advantages in terms of cost. On the one hand, no additional equipment, such as pipettes, sample vessels or reaction vessels, are required, and on the other hand, only the amount of measuring fluid required for the vessel "instant test" makes it possible to add labile components. Therefore, expensive reagents must be discarded. (e.g. due to lack of stability). As a result of the simplified operation, errors are avoided and therefore there is no need to perform repeated analyses. The simplified operation, which substantially reduces the possibility of errors, also allows persons without special training to perform the analysis and obtain accurate results. Therefore, these tests can be performed even where specially trained personnel are not available (eg, night work where medical technology employees are not available, and developing countries).
Having someone who is not specially trained to perform these container "instant tests" results in further economic savings. The present invention will be explained in more detail below with reference to the drawings. A glass or plastic container, which is schematically shown in the drawing, serves as the measuring container. The measurement container is filled to about half with reaction solution 1,
Its upper end is hermetically sealed by a cap 2. Since the cap 2 also serves as a handle for the container, the container will not be contaminated when it is gripped. The cap 2 has a pin 3 with a break setting position 4. When the pin 3 is laterally folded, a circular hole is formed in the top surface of the cap. Capillary tubes 5 and 6 can be inserted through this hole. The capillary tube 5 serves, for example, to aliquot a fluid sample.
The inner surface of the capillary tube 6 is coated with a solid reactant.
Instead of taking the form of a coating, the solid reactant may also be loosely packed within the capillary tube. When capillary tubes 5 and 6 are inserted and the container is shaken vigorously, the contents of capillary tube 6 and
The reactants in the capillary tube 6 are transferred to the reaction solution 1. The capillary tubes 5 and 6 are also located at the corners of the container by adhesive forces so that they do not interfere with the light path of the photometer. It is also conceivable that both capillaries 5 and 6 are coated inside with various reactants,
In this case, the capillary tube 5 is used at the same time for sample collection. Furthermore, there is also the possibility of operating with only one capillary. In this case, the capillary tube 5 used for sample collection simultaneously carries the reactants. For example, a commercially available device of the design shown in DE 23 38 206 can be used as a photometer. Example a Determination of hemoglobin by the hemoglobin cyanide method In this case, hemoglobin is oxidized to hemoglobin with potassium hexacyanoferrite(m), and hemoglobin is converted to hemiglopin cyanide with potassium cyanide. The quantity to be measured is the true absorbance produced by myglobin cyanide at a wavelength of 540-546 nm. Equipment: Spectrophotometer or filter photometer with a measuring wavelength of 540 or 54 m. Reagent: Plastic disposable measuring container with lid. Layer thickness: 1.00 skin; content: 1.25 reagent with the following composition: potassium hexacyanoferrite(m) 0.8
hmol/10 phosphate buffer [PH7.2 or 6.8
]2. Shoes mol/10 Sodium chloride 1.8 hmol/
1. Detergent (e.g. saponin) 0.05%. A graduated glass capillary tube with a capacity of 5 mu (length: 32 sides: inner diameter 0.446 ribs) whose interior is coated with KCN 70 g. Method: The breaking point 4 of the lid 2 of a plastic disposable measuring container is broken by means of a pin 3 provided for breaking. Five volumes of blood are taken, for example, from a blood-containing sample container using a volumetrically calibrated glass capillary tube or from a sample of blood from a fingertip. Any blood that may remain on the outside of the capillary is wiped away and the blood-filled capillary 5 is placed into the container. The hole formed in the lid 2 is sealed with an adhesive label. Blood is mixed with reagent 1 by shaking the measuring container, taking care to handle the measuring container only by the lid and bottom to avoid contaminating the walls (measuring surface) of the container. Reaction time (pH 7.2
After 3 minutes at pH 6.8 and 1 minute at pH 6.8), the true absorbance obtained by the analysis is compared with the true absorbance obtained by the photometer from an unused plastic disposable measuring container (= reagent control value). obtain. The resulting amount of hemoglobin in the blood is obtained as the number of grams of hemoglobin per 100 blood by multiplying the vacuum luminous intensity by 36.8. Preparation of capillary tubes coated inside with KCN Dissolve 9 parts of KCN in 5 parts of a suitable solvent (eg methanol or ethanol). The capillary tube 5 is filled by dipping into this solution. The solvent is removed under slight vacuum so that the KCN is evenly distributed on the inner surface of the capillary. When stored in a dry state. The contents of these capillaries are stable for at least 12 months. If the plastic disposable measuring container is packaged in a water vapor-impermeable manner (for example sealed with deep-drawn aluminum foil), its contents will also be stable for at least 12 months at a given volume. The measurement range and measurement sensitivity completely correspond to the reference method shown in the German Provisional PIN Standard No. 1. The accuracy and precision of the measurements were checked by several analyzes of control blood (Cou1 with 4 r-Counters).
C, CH60 of MerzandDade). As shown in Table 1 (Appendix), the same results were obtained for Co mountain ter
As when measuring hemoglobin using a hemocytometer manufactured by E1 electronics Gmb,
Obtained when measuring hemoglobin using the "instant" container of the invention. Regression line y=0.966 treated as 0.3570 (y=“immediate”
container; x = hemocytometer) and correlation coefficient r = +0.998
5 was calculated from the results of a comparative experiment conducted on 40 samples of human blood. According to these values there was a very close correlation between the two methods. B Determination of bilirubin using 2,5-dichlorophenyldiazonium salt In this determination, total bilirubin is combined with 2,5-dichlorophenyldiazonium chloride to form azobilirubin which reacts. Indirect bilirubin is released by detergents. The quantity to be measured is the true absorbance obtained with azobilirubin at the wavelength m. Equipment: Spectrophotometer or filter photometer with a measurement wavelength of 54 m. Reagent: Plastic disposable measuring container with lid. Layer thickness: 1.00 skin; Contents: Reagent 1 with the following composition
．． 25': Hydrochloric acid 0.1N Cleaning agent I% 2,6-dichlorophenyldiazonium chloride coated inside glass with capacity scale of 20 1 (length 3 shipboard; inner diameter 1.302 side) capillary tube. Glass capillary tube with volume scale (50 mu) for sample collection. Method A breaking point 4 on the lid 2 of a plastic disposable measuring container is broken by a breaking pin 3. 50 liters of sample, for example serum or blood*, are placed into a measuring container using a volumetric glass capillary tube 5. After sealing the hole in the diamond with an adhesive label, the sample is mixed with the reagent by gently shaking the measurement container. At this point, care must be taken not to touch the measuring surface. True absorbance obtained for the measuring container (E. = sample control value)
is measured with a photometer. Next, put the glass capillary tube 6 containing 2,5-dichlorophenyldiazonium chloride into the measuring container, and hold the container until the 2,5-dichlorophenyldiazonium chloride is dissolved (about 15 seconds). Shake lightly. After the reaction time has elapsed (at least 10 minutes at room temperature)
, the true absorbance (E2) for analysis is measured with a photometer. The resulting concentration of total bilirubin present is determined by subtracting the sample control value (E,) from the analytical value (E2) and dividing this difference by 44. It is obtained as milligrams per 100 samples (serum or blood) by sampling. 2. Preparation of a capillary tube whose interior was coated with 5-dichlorophenyldiazonium chloride 2.5-Dichlorophenyldiazonium chloride 125:9 was coated with a suitable fluid (
For example, suspend in 10 ml of ether). A 20 mu volume graduated glass capillary is immersed and filled with this solution. The fluid is removed under slight vacuum. In this way 2
,5-dichlorophenyldiazonium chloride is uniformly distributed over the inner surface of the capillary. The contents of these capillaries, if stored in a dry and cool place, will contain at least
Stable for 2 months. If the plastic disposable measuring container is packaged in such a way that it is impermeable to water vapor (
By sealing, for example with deep-drawn aluminum foil), these contents are also stable for at least 12 months in a given volume. The range and degree of sensitivity of the measurements obtained correspond to the method presented by Wahle Tsumugi [3]. The accuracy and precision of the measurements were checked by analyzing control sera several times. Measuring the total bilirubin concentration using the ready-to-use container gave results that were well comparable to those obtained by measuring total bilirubin by the method described by Wahl et al. [3]. Regression line 4 = 0.939 lines 0.1
804 (y = immediate container; × = test package, Boe
Hringer Mannheim's bilirubin DPD method) and the correlation coefficient r=10.9931 are calculated from the results of a comparative experiment performed on 40 samples. According to these values, there was a very close correlation between the two methods. Enzymatic cholesterol determination using cholesterol esterase, cholesterol oxidase, peroxidase and a subsequent color reaction by the method according to C. Trinder. In this method, cholesterol esters are linked by cholesterol esterase. Cholesterol is oxidized by cholesterol oxidase to form Δ4 cholestenone. The day 202 produced in this way is converted into phenol by the peracid enzyme.
and 4-aminophenazone to form the star-colored component 4-(p-benzoquinone-monoimino)-phenazone. The quantity to be measured is the true absorbance produced by 4-(p-benzoquinone-monoimino)phenazone at a wavelength of 546 nm. Equipment: Absorption photometer and filter photometer with a measurement wavelength of 54 meters. Reagent: Plastic disposable measuring container with diamond. Layer thickness: 1.00 mm; content: 1.5 mm with reagents having the following composition: potassium phosphate buffer (pH 7.7) 0
．． 4 mmol/1. Phenol 1 hmol/1. Methanol 1.8mm. 1/10 Hydroxypolyethoxydodecane 0.4%. 20 x 1 volumetric glass capillary tube (length 32 side, inner diameter 0.892) coated on the inner surface with 4-aminophenazone. Graduated glass capillary tube (20r1) for sample collection. Enzyme mixture in 3.2 M ammonium sulfate solution consisting of 20 U/' of cholesterol esterase, 6 U/' of cholesterol oxidase, and 4 U/' of peroxidase. Method: Break the break setting point on the lid of the plastic disposable measuring container with a break pin. 20 μ of the enzyme mixture are placed in a measuring vessel using a volumetric glass capillary whose inside surface is coated with 4-aminophenazone and mixed with the lightly shaken contents. In this way a complete reagent for cholesterol determination is created.
The true absorbance (E,=reagent reference value) given by the measuring container is measured with a photometer. Next, reagent 20〃1(
Serum or blood bran) is placed in a measuring container using a volumetric glass capillary tube, then shaken briefly and mixed with the reagent. After the reaction time has elapsed (at least 10 minutes at room temperature), the true absorbance (E2) for analysis is determined in a photometer. The resulting cholesterol concentration is determined as milligrams per sample (serum or blood acid) per sample (serum or blood acid) by subtracting the reagent control value (E,) from the analytical value (E2) and multiplying this difference by 652. be done. Preparation of a capillary tube whose interior is coated with 4-aminophenazone 457 parts of 4-aminophenazone are dissolved in a suitable fluid (eg dichloromethane or benzene) 20 parts. 2
A glass capillary tube with a 0 mu volume is immersed and filled with this solution. The fluid is removed under slight vacuum at 60 oo.
In this way, 4-aminophenazone is uniformly distributed on the inner surface of the capillary tube. The contents of these capillaries are stable for at least 12 months if stored in a dry, cool place. If the plastic disposable measuring container is made impermeable to water vapor, for example by sealing with deep-drawn aluminum foil, the contents are stable at a given volume for at least 12 months. The enzyme mixture is stable for at least 12 months if stored in a glass container at 14°C. The measurement range and the measurement sensitivity correspond to the method presented in Roschlau et al. [5]. The accuracy and precision of the measurements were checked by multiple analyzes of control sera. When measuring cholesterol using an immediate container, Rosc
Results were obtained that were sufficiently comparable to the results of cholesterol measurement by the method according to Hlau et al. [5], and in this case, the
The test package cholesterol CHOD-PAP method according to the rin trout rMan skin eim was used. Regression line y = 0.990 balls 12.7 I (r = immediate container; x
= Cholesterol CHOD-PAP Method Test Package, Bthrjn Bag rMan blood eim) was calculated from the results of comparative experiments performed on 40 samples, and the correlation coefficient was r = +0. The scale was 85. According to these values, a close correlation is formed between the two methods. Enzymatic determination of blood sugar using glucose oxidase, peroxidase and a subsequent star color reaction according to D. Trinder (example for multiple sensitive components in capillaries) By this method glucose is oxidized to gluconic acid by glucose oxidase. . The 2Q thus produced is reacted with 4-aminophenazone and phenol using peroxidase to form the color component 4-(p-penzoquinone-monimino)-phenazone. The quantity to be measured is the vacuum luminous intensity given by 4-(p-benzoquinone-monoamino)-phenazone at a wavelength of 54 m. Equipment: spectrophotometer or filter photometer with a measuring wavelength of 54 m. Reagent: Plastic disposable measuring container with lid. Layer thickness: 1 arc Contents: 1.5' of reagent with the following composition: phosphate buffer solution pH 7.0) 0.1 mmol/1. Phenol 4mmol no. A glass capillary tube with a capacity scale of 10 mm (length: 10 bars; inner wall: 1.12 sides) whose inner surface was coated with a mixture consisting of glucose oxidase, peracid enzyme, and 4-aminophenazone. A glass capillary tube with a capacity scale of 10 to 1 for sample collection. Method A break setting point on the lid of a plastic disposable measuring container is broken by a pin provided for breaking. A complete reagent is prepared by placing a volumetrically calibrated glass capillary containing a mixture of glucose oxidase, peroxidase and 4-aminophenazone in a measuring vessel.
Immediately after this, sample (e.g. serum or blood count) lo
w1 is placed in a measuring container using a volumetrically graduated glass capillary tube. The contents of the capillary tube are mixed with the contents of the measuring container by gentle shaking. Then the true absorbance given by the measuring container (E, = reagent control value and sample control value)
is measured by a photometer. After a reaction time (at least 2 minutes at room temperature), the true absorbance of the analysis (E2) is measured in a photometer. The resulting glucose concentration is calculated by subtracting the reagent control value and sample control value (E,) from the analytical value (E2).
By subtracting this difference from , and multiplying this difference by 597, you get the number of milligrams per 100 samples (serum or blood). Preparation of capillary tubes coated inside with glucose oxidase, peroxidase and 4-aminophenazone Glucose oxidase (purity 0100 U/9
) 500 evenings, peroxidase (purity DIOOU/9) 5
0.9 and 4-aminophenazone 520.9 are suspended in a suitable solvent (eg acetone or dichloroethane). A 10 mu volume graduated glass capillary is immersed and filled with this suspension. The fluid is removed under slight vacuum so that the enzyme and 4-aminophenazone are evenly distributed on the inner surface of the capillary. The contents of these capillaries are stable for at least 15 months when stored in a dry, cool place. If the plastic disposable measuring container is packaged so as to be impermeable to water vapor, for example by sealing with deep-drawn aluminum foil, the contents are stable at a given volume for at least 15 months. The measurement is linear over a range of 50 to 400 m9 of glucose per 100 m9 of sample. At high glucose concentrations the sample must be diluted. The accuracy and precision of the measurements are checked by analyzing control semen several times. Measuring Glucose Using Immediate Containers Boehringer Mannheim for Measuring Glucose
Results were obtained that were well comparable to those obtained using the automated packing glucose GOD-PAP method. Regression line y = 0.982 grade 0.7491 (y = immediate container; x = automated glucose GOD-PAP packaging method;
song ehrin trout rMan blood eim) and correlation coefficient r=
+0.9923 is calculated from the results of a comparative experiment performed on 40 samples. According to these values, there is a close relationship between the two methods. E Enzymatic measurement of blood sugar using the glucose dehydrogenase method (
Example for UV testing) By this method, BD-glucose is converted to D-gluconolactoso by glucose dehydrogenase. By doing so, NAD (nicotinamide adenine dinucleotide = coenzyme) is reduced to NADH2. The enzyme metarotating enzyme is the Q-D of 8-D-glucose.
- Promote formation from glucose. The force to be measured is the true absorbance versus NAD ratio at a wavelength of 34 or 36 meters. Equipment: Spectrophotometer or filter photometer with measurement wavelength of 340 or 36 mm. Reagent: Plastic disposable measuring container with lid. Layer thickness: 1c; Contents: reagent with the following composition: 1.
5': Phosphate buffer solution (Book 7.6) 0.1 mmol/
A glass capillary with a volume scale of 10 and 1 (length 1 stroke; inner diameter 1.12 ribs) whose inner surface was coated with a mixture consisting of 10 glucose dehydrogenases, metarotating enzymes and NAD. Glass capillary tube with 10 mu capacity scale for sample collection. Method This method determines reagent control values (necessary only once for each packing). For this purpose, a measuring vessel containing a buffer solution is measured against a measuring vessel containing a buffer solution and an internally coated capillary tube. The difference in true absorbance is the reagent control value (E.). sample (e.g. serum or 10 plates)
) is placed in an open measuring vessel and mixed with a buffer solution. The sample control value is then measured with a photometer (E,).
The capillary tube coated internally with glucose dehydrogenase, metarotating enzyme and NAD is then placed in the measuring vessel and the contents are mixed with the contents of the measuring vessel by gentle shaking. After the reaction time (10 minutes at room temperature) has elapsed, the true absorbance (E2) given by the analysis is measured in a photometer. As a result, the glucose concentration in milligrams per 100 samples is obtained by the following equation. Result = E2 - (E, 10 Eo) x 824 Fabrication of capillary tubes internally coated with glucose dehydrogenase, metarotating enzyme and NAD Glucose dehydrogenase 1
0 units, metarotating enzyme 3.8 units, and NAD 14.
9 of 6 is suspended in 10' of a suitable solvent (eg dichloromethane or dichloroethane). A 10 mu volume graduated glass capillary is immersed and filled with this suspension. The solvent was removed under slight vacuum to remove the enzyme and NAD.
Evenly distributed on the inner surface of the capillary tube. The contents of these capillaries are stable for at least 15 months when stored in a dry, cool place. The contents of the plastic disposable measuring container have a shelf life of at least 15 months when stored to be water vapor impermeable. The measurement range and degree of measurement sensitivity can be compared with the method presented by BaMuch [7]. The accuracy and precision of the measurements were checked by analyzing control sera several times. For the determination of glucose content using an extemporaneous container,
Results were obtained that were sufficiently comparable to those obtained with glucose measurements according to h[7], in which case Mark's test packing glucose (GIuc-DH method, UV test) was used. The regression line y=0.992 2.4120 [y=immediate container: 30×=Merckotest® Glucose] and the correlation coefficient r=+0.9941 are based on the comparative experiment performed on 40 samples. calculated from the results. According to these values, there is a close relationship between the two methods. Measurement of cholinesterase activity by the method according to FEI1man [kinetic method
Example for od)] According to the method described by EIlman et al. [8], cholinesterase opens acetylthiocholine to acetic acid and thiocholine. Thiocholine bromide reduces 5,5'-dithiobis-2-nitrobenzoic acid (DTNB) to 2-thio-2-nitrobenzoic acid, which is
It is measured by a photometer at a wavelength of 40 m. The quantity to be measured is the change in true absorbance given by 5-thio-2-nitrobenzoic acid at a wavelength of 405 nm. Apparatus: Spectrophotometer or filter photometer with measurement wavelength 405 and adjustable container holder overflow up to 2500. Reagent: Plastic disposable measuring container with lid. Layer thickness: 1.00 kg; Contents: 1.5 ml of reagent with the following composition: phosphate buffer (pH 7.2) 5 mol/
1. Saponin 0.1%. Acetylthiocholine diodide and DN
A glass capillary tube with a volume scale of 10 x 1 (length 1 x 1; inner diameter 1.12 side) whose inner surface is coated with a mixture with TB. Glass capillary tube with volume scale of 5〃1 for sample collection. Method Enzyme activity measurement is very strongly dependent on temperature, so the activity of cholinesterase must be measured at exactly 25 qo. For this purpose, a plastic disposable measuring container is heated to this temperature (for example by means of a temperature-adjustable container holder of a photometer). The complete reagent is formed by introduction of a capillary tube coated internally with azetylthiocholine diodide and TNB. Five mu of the sample (eg, serum, blood or whole blood) is then added using a volumetric capillary tube and mixed with the reagents by gentle shaking. The measurement container is placed in the photometer container, which is set at a temperature of 25°C. Initial true absorbance (E,
) is taken after 3 sands. E2 is measured exactly 6 seconds later. E3 is measured after 6 more sands. The difference in true absorbance per minute is obtained as the average value of E2-E and E3-E2. As a result, the activity of cholinesterase in units per milliliter (U/unit) is obtained by multiplying the difference in true absorbance per minute (ΔE/min) by 2.26. Preparation of capillary tubes coated with acetylthiocholine bromide and DTN separately 2.25% of acetylthiocholinedide and 150% of 5.5-dithiobis-2nitrobenzoic acid (DTNB) were dissolved in a suitable solvent (e.g. methanol). or ethanol) and suspend for 10 minutes. A 10-1 volume graduated glass capillary is filled by immersion into this suspension. The solvent is removed under slight vacuum so that the acetylthiocholine bromide and DTNB are uniformly distributed on the inner surface of the capillary. The contents of these capillaries are stable for at least 15 months when stored in a dry container. Plastic disposable measuring containers packaged to be impermeable to water vapor also last for at least 15 months. The measurement range and sensitivity are Frank

This corresponds to the method shown in [9]. Accuracy and precision were checked by analyzing control sera several times. When cholinesterase activity was measured using a ready-to-use container, results were obtained that were well comparable to the results of cholinesterase activity measured by EI1man et al. [8]. package is used).
The regression line y = 1.0030; Calculated from the results of According to these values, there is a close correlation between the two methods. Literature [1] C. STEFFEN: Photometer Best;mmu
ng derErythorZy nZahIAr
ZO. b. 5 (1959) 76-77 [2] L. J
ENDRASSI Ketal. :Bjochem. Z.
297 (1938) 81 [3] A. W. WAHLEF
ELDetal. :Scand. J. clin. Lab
．． Invest. Vol. 29, Suppl. 126(
1972) APStQCt11, 12 [4] Day. P.O.
PPERetal. : Biochem. Z. 291 (1937) 354 [5]
P. ROSCHLAUetal. :gh lnt. C
ongr. on CIin. Chemistry,T
oronto,. 1:975A is tr. The gunwale. 1 [6]”
P. TRINDER: Amn. CM. Biocbem.
6 (1969) 24 [7] D. BANAAUCHet
al. :Z. K1in. Chem. K1in. BiMh
em. 13 (1976) 101 [8] G. L. ELL
MANetal. :Biochem. Pharmaco
l. 7 (1961) Fence

[9] G. FRANK:Z'A
nalre. Chem. 279 (1976) 155 Brief description of the drawing The drawing schematically shows a container made of glass or plastic that serves as a measuring container. 1... Reaction solution, 2... Cap, 3... Pin, 4... Destruction setting position, 5, 6... Capillary tube.

Claims (1)

[Claims] 1. For use in an optical analysis device comprising a closed container containing a precisely prepared reaction solution and a capillary tube insertable into the reaction solution for dispensing a sample fluid into the solution. In an apparatus for the preparation of a sample fluid, the optical properties of the measurement fluid are such that the optical properties of the measurement fluid are such that at least one of the reactants required for the formation of the measurement fluid from the reaction solution is Device for producing a measuring fluid for the optical analysis device, characterized in that it is prepared by placing it as a solid phase in a capillary tube and/or in a second capillary tube that can be introduced into the container. 2. Apparatus according to paragraph 1, wherein the at least one reactant is applied to the inner wall of the capillary tube in finely divided form. 3. To measure hemoglobin concentration, the reaction solution in the container is phosphate-buffered hexacyanoferrous(III).
3. Apparatus according to claim 1 or 2, comprising a potassium acid/sodium chloride solution and wherein the capillary tube used for sample preparation is coated with potassium cyanide. 4. To measure bilirubin in blood, the reaction solution is
Said first capillary comprising 0.1N hydrochloric acid and, in addition to the capillary used for blood sample preparation, is provided with a second capillary coated with 2,5-dichlorophenyldiazonium chloride. or the device described in item 2. 5. For the measurement of cholesterol in serum or plasma,
The reaction solution comprises a phenol-methanol solution supplemented with hydroxypolyethoxide dodecane buffered with potassium phosphate, and a second capillary coated with 4-aminophenazone in addition to the capillary used to prepare the serum or plasma. 3. A device according to claim 1 or 2, wherein is provided for introducing an enzyme mixture consisting of cholesterol esterase, cholesterol oxidase and peroxidase. 6. For enzymatic blood glucose determination in serum or plasma by the method according to Trinder, the reaction solution contains phosphate-buffered phenol and is added to the capillary tube used to prepare the serum or plasma sample. In addition, glucose oxidase, peroxidase and 4-
3. A device according to claim 1 or 2, wherein a second capillary tube is provided coated with a mixture consisting of aminophenazone. 7. To carry out enzymatic blood glucose measurement by the glucose dehydrogenase method, the reaction solution comprises a phosphate buffer solution with a pH value of 7.6, and in addition to the capillary tube used for sample preparation, glucose dehydrogenase, 3. A device according to claim 1 or 2, wherein the second capillary tube is coated with a mixture consisting of metarotactic enzyme and nicotinamide adenine dinucleotide (NAD). 8 To measure cholinesterase activity by the method according to Ellman, the reaction solution was adjusted to a pH of 7.
．． In addition to the capillary tube used to prepare the sample, a second capillary tube containing a phosphate buffer solution of 2 and coated with a mixture consisting of acetylthiocholine iodide and dithiobis-2-nitrobenzoic acid is provided. 3. The device according to claim 1 or 2, wherein the device is made of: