JPH06324047A - Measuring apparatus for endotoxin - Google Patents

Abstract

PURPOSE:To automatically detect whether the test when the quantity of endotoxin is measured is valid and whether the preparation of products is proper, and to check necessary conditions for starting the test before the measurement is started thereby to completely avoid waste brought about if the measurement is tried again. CONSTITUTION:The apparatus is provided with a preserving means 5 for preserving at least the arrangement of kinds of samples of specimens and comparison specimens, means 9, 14 for measuring the reaction of a to-be-inspected sample which is a mixture of the specimen or comparison specimen with a reagent thereby generating a measuring value, means 6, 7, 8 for preserving conditions to execute calculations to the measuring value of the to-be-inspected sample, and means 10, 11 for inputting a predetermined data to each preserving means. Further, the apparatus has judging means 3, 4 and means 12, 13 for outputting the result of tests. Each judging means 3, 4 suitably combines the contents of the preserving means in accordance with the kind of a test, detecting necessary conditions for starting the test, performing calculations from the measuring value and the content of each preserving means and judging the validity of the test and whether the calculated value is proper.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an analyzer for performing endotoxin test on product formulations and the like.

[0002]

2. Description of the Related Art Endotoxin is a representative of a pyrogen (pyrogen), and endotoxin is produced by the use of endotoxin-contaminated blood, infusion, injection, or the use of medical instruments contaminated with endotoxin. It is known that when injected into a living body, it causes serious side effects such as strong fever and shock. Therefore,
In these product formulations and the like, it is essential to measure the amount of endotoxin in it and prevent its contamination.

In actual measurement of endotoxin, endotoxin in a sample must be detected at a picogram level in a normal environment where a large amount of endotoxin is present, and pollution from the environment is always a problem. For this reason, it is necessary to have a high level of skill in the measurement procedure, such as the horseshoe crab blood cell extract reagent used in the reaction (the horseshoe crab reagent) and the handling of the instrument, and it is necessary to constantly check whether the measurement was performed with the correct procedure without contamination. is there. Usually, for this purpose, a sample containing no endotoxin is simultaneously measured as a negative control, and it is confirmed that this sample shows only a reaction at a predetermined reaction rate or less. In addition, it has been confirmed that the correlation coefficient of the calibration curve prepared with the endotoxin standard sample containing the predetermined concentration of endotoxin is within the predetermined range (validity check of measurement operation).

The reaction between the horseshoe crab reagent and endotoxin is often affected (inhibited or promoted) by various chemical substances in the product formulation to be measured. You need to check if the measurements were made below. To do this, add a predetermined concentration of endotoxin to the sample, confirm that the reaction rate with the horseshoe crab reagent is within the predetermined range, and then determine the reaction conditions (generally the dilution rate of the drug to be the sample). It is common to carry out a preliminary test (check for promoting inhibition of test drug).

In daily tests, a negative control measurement is checked, a positive control is sometimes added with a certain amount of endotoxin, and an endotoxin is added to a sample under a predetermined reaction (dilution) condition (endotoxin spike). (Sample) The amount of endotoxin contained in various samples must be measured while always checking whether or not the appropriate conditions for the reaction are reproduced even in the measurement at that time (daily inspection check). In addition, the concentration that establishes the appropriate reaction conditions for each type of product formulation, the sensitivity required for the measurement, and the allowable standard value of the endotoxin concentration differ, so the correct conditions should be used when measuring many samples. A complicated procedure is required to check whether or not the measurement is performed in.

[0006] At present, there is commercially available a device for measuring the reaction between endotoxin and horseshoe crab reagent and determining the endotoxin concentration based on a calibration curve. For example, a synthetic substrate method using a microplate reader and a turbidimetric time method using a dedicated device (JP-A-61-11641). Although the endotoxin concentration can be determined by the measurement using these devices, the measurement operator manually uses the measured values to perform the above-mentioned validity check of the measurement operation, check inhibition test of the test drug, daily check, etc. Must be done in work.

Because of the high sensitivity of the reaction between endotoxin and horseshoe crab reagent, it is difficult to obtain the same absolute measurement value every time, and the comparison sample (negative control mentioned above) must be used for the evaluation of the validity of the measurement result and the calculation of the sample concentration. , Positive control, spiked sample, endotoxin standard sample, etc.). Due to such circumstances and reagents being extremely expensive, data for simultaneous measurement of many samples and sharing of comparative samples required for individual sample measurement to reduce the number of samples, as well as simultaneous comparison with each other Often improve the reproducibility of. When measuring such a large number of test samples, more check items are required, and more skill is required to obtain a correct result. In addition, the reaction between the sample and the horseshoe crab reagent is unstable, so if even one of the required check items has not been performed, not only re-measurement of that item but also the sample and comparison sample that require comparison at the same time. All measurements must be redone at the same time. For example, the sensitivity of the calibration curve is insufficient, or the dilution exceeds the MVD (maximum effective dilution ratio: maximum value of the dilution ratio that the reaction dilution conditions can be considered effective), or the comparative sample required for checking has not been measured. If the number of comparative samples does not meet the standard, it is necessary to redo not only the re-measurement of the defective sample but also the whole measurement.

[0008]

That is, in the prior art, particularly when measuring many samples at the same time, the measurement conditions and individual tests required for each sample (validity check of the measurement operation, test Checking the promotion of drug inhibition,
Setting of various conditions required for measurement, such as the setting conditions required for daily inspections, etc., is complicated and requires skill.
When the re-measurement occurs due to the lack of necessary conditions after the measurement, the re-measurement is required, resulting in a large amount of wasted labor and time, and an increase in various expenses.
This invention has been made to solve the above-mentioned problems, and automatically determines the effectiveness of the test by measuring the amount of endotoxin, and automatically determines the acceptance or rejection of the product formulation,
It is an object of the present invention to provide an endotoxin measuring device capable of completely checking the necessary conditions for starting a test before starting the measurement and completely eliminating the waste caused by re-performing the measurement.

[0009]

The above objects of the present invention are as follows.
A means for storing at least the sample type sequence of the sample and the comparative sample, a means for measuring the reaction of the test sample in which the sample or the comparative sample and the reagent are mixed and generating a measurement value, and an operation for the measurement value of the test sample A means for storing the conditions to be performed, a means for inputting predetermined data to each of the storage means, an appropriate combination of the contents of each of the storage means according to the type of test and a necessary condition for starting the test are determined, and the measured value And an endotoxin measurement device characterized by having means for performing calculation from the contents of each of the storage means and for judging the validity of the test and whether the calculated calculated value is acceptable or not, and means for outputting the result of the test. .

[0010]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS An embodiment of the present invention will be described below with reference to the drawings. The examples of the present invention are used as a quality control procedure for the endotoxin test in the US Food and Drug Administration (FDA: Food and D
For example, the guidelines issued by (rug Administration) are explained. According to this guideline, when conducting routine inspections of actual product formulations, the following shall be performed. As a sample, a product sample, and as a comparative sample, an endotoxin spike sample, an endotoxin standard sample, and a negative control are included at least twice. In addition, it is necessary to include endotoxin standard samples at 4 concentrations or more. An endotoxin spike sample is a product sample to which a known amount of endotoxin is added. If the negative control exceeds a certain concentration as a result of the measurement, or if the absolute value of the correlation coefficient of the linear regression equation of the endotoxin standard sample falls below a certain value, the test is judged to be invalid. For the actual product sample, the value obtained by subtracting the endotoxin concentration of the product sample from the endotoxin concentration of the endotoxin spike sample is within ± 25% of the added endotoxin concentration, and the endotoxin concentration of the product sample is limited by the product. If it is within the value, it passes the test.

FIG. 1 shows the construction of the endotoxin measuring apparatus of this embodiment. This endotoxin measuring device 1
Is roughly composed of a computer system 2 and a measuring device 14. Each device is connected by a cable capable of transmitting data as needed. Computer system 2
Is a central processing unit (CPU) 3, memory groups 4 to 9,
Input / output groups 10 to 13 are provided. Each device is connected via a bus and an interface.

The program memory 4 stores programs necessary for setting various parameters and operating the measuring device to acquire data. Measurement data memory 9
Stores measurement data obtained from the measuring device or data obtained by appropriately processing the measurement data. The calibration curve data memory 8 stores the calibration curve data for converting the measurement data into the endotoxin concentration. The keyboard 10 is used for inputting commands when setting conditions, entering a file name, designating an output format, and the like. The disk drive 11, display 12, and printer 13 are general-purpose.

The sample array table memory 5 stores a sample setting table. This is a table that stores, for each sample placed in the measuring device 14, an array of sample types such as a product sample, an endotoxin spike sample, an endotoxin standard sample, and a negative control. The standard value table memory 6 is
Parameter values according to the sample type (for example, endotoxin concentration value for endotoxin standard sample, dilution rate for product sample, added endotoxin concentration for endotoxin spike sample, etc.) , And stores the standard value and the like for the determination of the effectiveness of the test and the acceptance / rejection of the test drug. These include, for example, the effective limit value of the calibration curve correlation coefficient and the effective limit value of the endotoxin addition recovery rate. The parameter value corresponding to the sample type can be stored in the memory 5 as data associated with the sample array. The drug information memory 7 stores information on the drug to be tested. This includes, for example, the name of the drug, endotoxin limits, potency, etc. The conditions stored in the memories 6, 7, and 8 determine the conditions for performing calculations on the measured values of the test sample. In addition to the conditions described here, the conditions for the calculation may be integrated into one memory, or may be divided and stored in a memory that is different from the one described here. Also, once these information contents are set, they can be shared information that can be used in tests other than daily inspections.

The measuring device 14 is a device for optically measuring the amount of endotoxin (colorimetrically or turbidimetrically) and comprises a sample holding part, a light emitting part and a light receiving part. In order to measure a plurality of samples in parallel, it may have a plurality of sample holders, light emitting units, and light receiving units. This device 14 can generate a measured value by a synthetic substrate method, a turbidimetric time method, or the like. The conventional apparatus does not include the sample array table memory 5, the standard value table memory 6, and the drug information memory 7, and naturally, processing other than calibration curve calculation is impossible.

Next, the operation of the apparatus of the embodiment will be described with reference to the flowchart of FIG. In step 1, the operator inputs the sample sequence data from the keyboard. Alternatively, it is possible to input the data which has been set and saved in advance by reading it from an external storage device. As the external storage device, for example, a floppy disk, a hard disk, a magneto-optical disk, a memory card or the like can be used. This setting is stored in the memory 5. If necessary, in step 2, input the previously measured calibration curve data from an external storage device,
It is stored in the memory 8. In step 3, standard value data and the like for the test validity determination and the product sample pass / fail determination are input from the keyboard and stored in the memory 6. Input to the memory 6 can be similarly performed through an external storage device.

In step 4, the test drug name, titer,
Information regarding the test drug such as the endotoxin allowable limit value is input and stored in the memory 7. Instead of manually inputting each item of drug information, the name of the drug can be specified and input from a database file on an external storage device in which these items are registered in advance. The memories 5, 6, 7, and 8 may be RAM inside the computer or external storage devices. Above, Step 1
The execution of the inputs 4 to 4 can be performed in various ways depending on the program, such as further dividing individual inputs or collectively performing a plurality of inputs.

In step 51, the type of test such as daily inspection is designated from the keyboard, etc., and in step 52, it is checked from the set sample sequence data and calculation conditions whether or not the necessary conditions for starting the desired test are satisfied. To do. Prior to the check, a new reference value for the check may be created from the information individually set in steps 1 to 4 according to the purpose of the test. For example, the MVD may be calculated for each sample from the minimum endotoxin concentration of the endotoxin standard sample in the standard value table memory 6 and the endotoxin limit value of the test drug in the drug information memory 7. Check the necessary conditions to start the test with the CPU
According to the program. As the contents of the check, for example, a negative control, an endotoxin standard sample, a product sample, a check on the number of samples / repetition number of the endotoxin spike sample, a check on whether the dilution ratio of the product sample does not exceed the MVD, etc. are performed. If the necessary condition is not satisfied, the process cannot be moved to the next step until it is satisfied, and the process is repeated from the first step. At this time, it is also possible to give a warning by notifying the contents of the inappropriate condition setting.

Only if the requirements are met in step 52 can the actual measurements be taken. That is, since the measurement is not started under the wrong condition setting, the situation of redoing all the measurements is not caused.
In the present invention, the information in the memories 5, 6, 7, and 8 is shared as individual information independent of the type of test, and is appropriately selected and combined according to the purpose of the test and processed. In step 6, the sample is measured by the measuring device 14. The measurement data is stored in the memory 9. The memory 9 can be placed in the measuring device 14 or in an external storage device.

When the measurement of all the samples is completed, the process proceeds to step 7, and if the endotoxin standard sample is set in the sample sequence table memory, the calibration curve is calculated from the measurement data of the endotoxin standard sample, The calibration curve data is stored in the memory 8. Then, the process proceeds to step 8, and the endotoxin concentration values of all the samples are calculated from the measurement data using the calibration curve data (the data read in step 2 or the data calculated in step 7). In step 9, memory 5,
Based on the information stored in 6, 7, and 8, the calculation of the output data and the calculation of the standard matching value for matching with the standard value are executed for each sample. Steps 7, 8 and 9 are performed by the CPU according to a program.

In step 10, the validity of the test is determined from the processed calculated value. It is effective if the calculated concentration value is less than the standard value for the negative control sample. It is effective if the absolute value of the correlation coefficient of the linear regression is greater than or equal to the standard value for the calibration curve data. In step 11, the calculated value is processed to determine whether the test drug is acceptable or not. First, it is checked whether the endotoxin addition recovery rate of the endotoxin spiked sample calculated in step 9 is within the range of the standard value stored in the memory 6. Further, it is checked whether or not the concentration value of the product sample calculated in step 9 is less than or equal to the endotoxin allowable limit value for the drug stored in the memory 7.
Only when the above two conditions are satisfied, the test drug is judged to pass. The CPU determines the validity of the test in step 10 and the pass / fail determination in step 11 according to a program.

Finally, in step 12, a report of the test result including the validity of the test and the pass / fail judgment of the product drug is output to the display 12 or the printer 13.
Alternatively, the data can be output to an external storage device or the data can be transferred through a communication line. Also in the validity check test of the measurement operation and the inhibition promotion check test of the test drug, in steps 51 and 52, the setting contents of the sample sequence table 5, the standard value table 6, the drug information memory 7, and the calibration curve data memory 8 are used. By checking the necessary conditions for starting the test set for each test, it is possible to avoid the situation in which the measurement must be started with the wrong setting and the measurement must be performed again. In addition, when performing multiple tests at the same time, it is possible to check the necessary conditions for starting the test by designating simultaneous execution of the multiple tests in step 51, and to share information between the tests (for example, the sequence of the comparison sample). , Standard values, etc.) and measurement data of comparative samples and the like can be combined to determine the effectiveness of the test set for each test with the minimum required sample.

Here, the present invention has been described by taking the FDA guideline as an example. However, according to the apparatus of the present invention, the contents such as the necessary condition check for starting a test included in the program memory and the memories 5, 6, 7 are stored. By simply changing the input contents, various test methods and standards other than those specified in these guidelines can be easily and flexibly supported. For example, as shown in FIG. 3, by adding a memory 15 that stores a group of subroutines programmed with different test procedures, even when tests of different methods are simultaneously performed, the necessary conditions for starting the test are checked, and after the measurement, It will be a system that can check the validity of the test and judge the pass / fail of the sample. With a system that simultaneously measures samples that perform different tests in this way, a negative control, an endotoxin standard sample, and other comparative samples are shared in all tests to reduce the number of samples and save the use of expensive horseshoe crab reagents. It is possible to

[0023]

As described above, until now, there was no endotoxin measuring device that automatically executes the effectiveness of the test and whether the product formulation passed or failed and outputs the result.
According to the present invention, an endotoxin measurement device capable of not only simply measuring the amount of endotoxin but also automatically determining the effectiveness of the test and the acceptance of the product formulation and outputting a report of the result. Can be provided. Further, since the necessary condition for starting the test can be checked before starting the measurement, it is possible to completely eliminate the waste caused by the re-measurement. Furthermore, since a comparative sample can be shared by simultaneously executing a plurality of tests, the amount of expensive horseshoe crab reagent used can be reduced.

[Brief description of drawings]

FIG. 1 is a configuration diagram of an endotoxin measurement device according to an embodiment of the present invention.

FIG. 2 is a flowchart illustrating the operation of the device of FIG.

FIG. 3 is a configuration diagram of an endotoxin measurement device according to another embodiment of the present invention.

[Explanation of symbols]

 1 Endotoxin Measuring Device 2 Computer System 3 Central Processing Unit (CPU) 4 Program Memory 5 Sample Sequence Table Memory 6 Standard Value Table Memory 7 Drug Information Memory 8 Calibration Curve Data Memory 9 Measurement Data Memory 10 Keyboard 11 Disk Drive 12 Display 13 Printer 14 Measuring device

Claims (1)

[Claims] 1. A means for storing at least an array of sample types of a sample and a comparative sample, a means for measuring a reaction of a test sample in which a sample or a comparative sample and a reagent are mixed, and generating a measurement value, a measurement value of the test sample Means for storing the conditions for performing the calculation, means for inputting predetermined data to each of the storage means,
Depending on the type of test, the contents of each of the storage means are appropriately combined and the necessary condition for starting the test is determined, and calculation is performed from the measured value and the content of each of the storage means and the effectiveness and the test are calculated. Means for judging whether the calculated value is acceptable or not,
An endotoxin measuring device comprising: means for outputting a test result.