JP2021124455A - Dissolution method of drying reagent - Google Patents

Abstract

The present invention provides a method for preventing defective dissolution of a dry reagent by checking the progress of dissolution in real time during the dissolution of the dry reagent. [Solution] A step of adding a solvent to a dry reagent placed in a container, and starting detection of fluorescence intensity by aiming the effective diameter of excitation light and fluorescence of a fluorescence detector so as to cover all of the dry reagent. and stirring the container until the detected fluorescence intensity reaches an arbitrary threshold value. [Selection diagram] Figure 1

Description

The present invention relates to a method for dissolving a drying reagent.

Dry reagents are often used in automatic analyzers that analyze components in biological samples by mixing and reacting biological samples with reagents. The drying reagent needs to be dissolved in a solvent, and is usually carried out at a constant stirring time and temperature (see, for example, Patent Document 1). If the drying reagent is not sufficiently dissolved, it may be erroneously determined due to a decrease in the reactivity of the reagent, but it was only visually confirmed whether or not the drying reagent was sufficiently dissolved.

WO2017 / 149940 Pamphlet

By confirming the progress of dissolution in real time during the dissolution of the drying reagent, a method for preventing the dissolution failure of the drying reagent is provided.

In order to solve the above problems, the present inventors have reached the present invention as a result of repeated diligent studies.
That is, the present invention detects the fluorescence intensity by aiming at the step of adding a solvent to the drying reagent arranged in the container and aiming so that the excitation light of the fluorescence detector and the effective diameter of fluorescence cover all the drying reagents. A method for dissolving a drying reagent, comprising a step of starting and a step of stirring the inside of the container until the detected fluorescence intensity reaches an arbitrary threshold value.

Hereinafter, the present invention will be described in detail.

The drying reagent is a reagent from which water has been removed by freeze-drying, air-drying, vacuum-drying, or the like a liquid reagent. Examples of nucleic acid amplification reagents include, but are not limited to, dry reagents containing enzymes, substrates, primers, and fluorescent labels that specifically react with augmented by-products. The shape may be granular, powdery, or thin film, but the thin film is particularly preferable.

The container in which the drying reagent is placed is made of plastic, which is easy to dispose of, although the material, size, and shape are not particularly limited as long as it does not affect the dissolution reaction of the reagent or the nucleic acid amplification of the sample. Container is preferable.

As the solvent used to dissolve the drying reagent, purified water to which the measurement sample is added is used. Depending on the reaction between the drying reagent and the measurement sample, an acid, a salt, or a surfactant may be further added to the solvent as appropriate.

After adding the solvent to the drying reagent placed in the container, the fluorescence intensity is detected by aiming so that the excitation light of the fluorescence detector and the effective diameter of fluorescence cover all the drying reagents. Usually, the drying reagent has a heavier specific gravity than the solvent, so if the container is large enough, the drying reagent is submerged in the inner bottom of the container.

As the fluorescence detector to be used, there is no problem as long as it is a general device including an excitation light irradiation means for irradiating the sample with excitation light and a fluorescence detection means for detecting the fluorescence emitted from the sample, but there is no problem with 3 excitation 3 Fluorescence detectors are particularly suitable. The effective diameters of the excitation light and the fluorescence refer to the effective diameters of the lenses provided by the excitation light irradiation means and the fluorescence detection means (see JIS B 7095: 1997). That is, a circle having an area equal to the cross-sectional area perpendicular to the optical axis of a bundle of parallel rays that should exit from an infinite object point on the optical axis of the lens and pass through a lens having an aperture corresponding to a given aperture scale. Refers to the diameter of.

The position of the fluorescence detector is not particularly limited as long as the effective diameter of the excitation light and fluorescence covers all the drying reagents, and an incubator for nucleic acid amplification and a fluorescence detector can be integrated. Is.

When the fluorescence intensity is detected by aiming so that the excitation light of the fluorescence detector and the effective diameter of fluorescence cover all the drying reagents, the fluorescence intensity is highest at the stage where dissolution has not started (drying reagent only), and the drying reagent. Is evenly dissolved in the solvent, the fluorescence intensity is lowest. That is, as the dissolution of the drying reagent progresses, the fluorescence intensity gradually decreases.

Using the above principle, an appropriate fluorescence intensity is set as a threshold value. If the fluorescence intensity reaches an arbitrary threshold value, it can be determined that the drying reagent is evenly dissolved in the solvent, and at that point, stirring for promoting dissolution may be stopped. The stirring method can be exemplified by a method using a stirring blade, a method of vibrating the container horizontally or vertically, a suction discharge by a dispensing mechanism, and the like, but there is no particular limitation.

Prevents poor dissolution of the drying reagent in the container, does not perform unnecessary stirring, and contributes to overall time reduction.

It is the schematic which shows the dissolution method of the drying reagent of this invention. It is a figure which showed the result of Example 1. FIG.

Hereinafter, specific embodiments will be described, but those skilled in the art will appreciate that the present invention is not limited to this example and that various modifications can be made within the scope of the invention described in the claims. It is easy to understand.

Example 1
Using the fluorescence detector of the present invention and the stirring mechanism shown in FIG. 1, it was verified whether the progress of dissolution of the drying reagent could be measured by fluorescence in real time. A colorless and transparent polypropylene container was used as the sample container 10, a nucleic acid probe modified with a fluorescent dye was used as the drying reagent 11, and a solvent containing purified water as a main component was used as the solvent 12. As the excitation light irradiation unit 14 and the fluorescence detection unit 15, three sets of excitation light 450 nm fluorescence 495 nm, excitation light 500 nm fluorescence 545 nm, and excitation light 590 nm fluorescence 645 nm were used. The three excitation light irradiation units 14 were turned on in order (2.5 sec × 3, 2.5 sec off), and the fluorescence intensity of the corresponding fluorescence detection unit 15 was measured. The solvent 12 was discharged into the sample container 10 by the dispensing mechanism 13, and then stirring was performed by suction discharge.

The results are shown in FIG. Before discharging the solvent 12, all the drying reagents were covered within the effective diameter of the fluorescence detector, so that all three types of fluorescence detectors received high fluorescence intensity. The fluorescence intensity gradually decreased for 40 seconds after discharging the solvent 12. It can be seen that the drying reagent is temporally dissolved in a solvent outside the effective diameter. After 40 seconds, the fluorescence intensity became constant. It can be seen that all the drying reagents were dissolved in the solvent.

10: Sample container 11: Drying reagent 12: Solvent 13: Dispensing mechanism (stirring mechanism)
14: Excitation light irradiation unit 14a: Excitation light source 14b: Optical filter (excitation light side)
14c: Effective diameter of excitation light 15: Fluorescence detection unit 15a: Light receiving element 15b: Optical filter (fluorescence side)
15c: Fluorescence effective diameter

Claims (4)

The process of adding a solvent to the drying reagent placed in the container,
A step of aiming the excitation light of the fluorescence detector and the effective diameter of the fluorescence so as to cover all the drying reagents, and a step of starting the detection of the fluorescence intensity.
The step of stirring the inside of the container until the detected fluorescence intensity reaches an arbitrary threshold value, and
A method for dissolving a drying reagent comprising. The method according to claim 1, wherein the fluorescence detector is a 3-excitation 3-fluorescence detector. The method according to claim 1 or 2, wherein the shape of the drying reagent is a thin film. The method according to any one of claims 1 to 3, wherein the drying reagent is a nucleic acid amplification reagent.

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