JP2019012035A - Kit for thin layer chromatography, developer for thin layer chromatography, and thin layer chromatography - Google Patents

Abstract

To provide a kit capable of more accurately detecting 8OHdG in urine than conventional arts.SOLUTION: A kit for thin layer chromatography comprises: an introduction part into which a sample containing 8-Oxo-2'-deoxyguanosine and impurities is introduced; developer for developing the 8-Oxo-2'-deoxyguanosine and impurities in the sample; a detection unit configured to detect an amount or concentration of the 8-Oxo-2'-deoxyguanosine contained in solution obtained by developing the sample with the developer, in which a speed at which the 8-Oxo-2'-deoxyguanosine is developed with the developer is faster than a speed at which the impurities are developed with the developer.SELECTED DRAWING: Figure 1

Description

  The present invention relates to a kit for thin layer chromatography, a developing solution for thin layer chromatography, and thin layer chromatography.

  8-Oxo-2'-deoxyguanosine (8 oxo 2 'deoxyguanosine, hereinafter abbreviated as 8OHdG) is known as a substance that reflects the effect of active oxygen on the living body, and is a widely used oxidative stress marker one of. It is thought that measuring the amount of 8OHdG produced in the body leads to understanding of the disease state and disease prevention. Since 8OHdG is a chemically stable substance that is excreted in the urine without undergoing secondary metabolism, it is possible to measure oxidative stress markers non-invasively by measuring the amount of 8OHdG in the urine. It is.

  Patent Document 1 discloses a system for determining an oxidative stress state using an antibody that specifically reacts with 8OHdG. Specifically, a sample is added to a test kit in which a conjugate of gold colloid bound to an antibody that specifically reacts with 8OHdG is immobilized, and the amount of the conjugate that has not bound to 8OHdG is determined by color. By doing so, detection of 8OHdG is performed. However, since the system disclosed in Patent Document 1 does not use a solution for developing a sample, it is difficult for the conjugate to reach the position where the amount of the conjugate is determined, and there is a possibility that accurate determination cannot be made.

  Patent Document 2 discloses a method and apparatus for quantifying 8OHdG concentrated by pretreatment by high performance liquid chromatography (HPLC). Patent Document 1 discloses performing separation and concentration using a solution obtained by concentrating 8OHdG using silica gel or ethanol as pretreatment for quantitative determination by HPLC.

JP 2010-156039 A JP 2007-121271 A

  Here, the present inventors have found that there is a problem with the method of Patent Document 2. In Patent Document 2, ethanol is used for the separation and concentration of 8OHdG. However, when the sample is urine, it is difficult to separate 8OHdG and uric acid sufficiently, and it is difficult to accurately detect 8OHdG. It was.

  This invention is made | formed in view of the said subject, and it aims at providing the kit which can detect 8OHdG in urine more accurately than before.

  The kit for thin-layer chromatography according to the present invention includes an introduction part into which a sample containing 8 oxo 2 ′ deoxyguanosine and a contaminant is introduced, and among the samples, the 8 oxo 2 ′ deoxyguanosine and the contaminant. A thin layer chromatography comprising: a developing solution for separation; and a detection unit for detecting the amount or concentration of the 8oxo-2′deoxyguanosine contained in a solution obtained by developing the sample with the developing solution. The development kit is characterized in that the development rate of the 8 oxo-2′deoxyguanosine by the developing solution is faster than the development rate of the contaminants by the developing solution.

  Another thin-layer chromatography kit according to the present invention includes an introduction part into which a sample containing a detection target is introduced, a developing solution for separating the detection target among the samples, and the sample is the And a detection unit for detecting the amount or concentration of the detection target contained in the solution developed by the developing solution, wherein the developing solution is methanol, ethanol , At least one of propanol and isopropanol, and at least one of potassium hydroxide and sodium hydroxide.

  The developing solution for thin-layer chromatography according to the present invention contains at least one of alcohol and ether and an inorganic electrolyte.

  The thin layer chromatography according to the present invention is a thin layer chromatography for detecting 8 oxo 2 ′ deoxyguanosine, introducing a sample containing the 8 oxo 2 ′ deoxy guanosine and impurities, and developing the sample. A step of separating and extracting 8OHdG from the sample by developing a liquid, and a step of detecting the amount or concentration of the separated 8 oxo-2′deoxyguanosine.

  According to the kit for thin-layer chromatography according to the present invention, 8OHdG in urine can be detected with higher accuracy than in the past.

The figure for demonstrating the kit for thin layer chromatography to embodiment of this invention. The figure for demonstrating the method to detect 8OHdG using the kit for thin layer chromatography which concerns on embodiment of this invention. The graph showing the result of having quantified 8OHdG using the kit for thin layer chromatography which concerns on embodiment to this invention.

  Hereinafter, although embodiment of this invention is described, this invention is not limited to these.

<Kit for thin layer chromatography>
An example of a detection kit for thin layer chromatography will be described in this embodiment with reference to FIG. The kit according to this embodiment includes an introduction unit 102 for introducing a sample onto a support 104, a separation unit 103 for separating a detection target in the sample and at least a part of other substances, A detection unit 106 is provided for detecting the amount or concentration of the detection target. In the present embodiment, an introduction unit 102 and a detection unit 106 are provided in a part of the separation unit 103.

  A developing liquid L, which will be described later, is introduced from the developing liquid introducing unit 105, and the sample 101 introduced into the introducing unit 102 starts moving toward the detecting unit 106 by the developing liquid L. FIG. 1 shows an example in which the developing solution is held in the holding unit 110, but the invention is not limited thereto, and the developing solution may be introduced into the developing solution introduction unit 105 from the outside.

  The speed of the sample moving through the separation unit 103 varies depending on the physical properties of each component in the sample. When the developing solution reaches the detection unit 106, each component in the sample is a region from the separation unit 103 to the detection unit 106. Distributed in different places. The detection illumination 107 is irradiated to the detection target 109 to be quantified, and the detection target is quantitatively detected by the signal receiving unit 108. In the present specification, it is desirable that the quantitative detection of the detection target is strictly known in terms of the amount and concentration. For example, A (0 to 10%) B (10% to 30%) C (30% to 50) %) D (50% or more) is a concept including the fact that it belongs to one of a plurality of ranges. Or the information regarding the quantity and density | concentration of a detection target object can be acquired using the kit which concerns on this embodiment.

  As described above, the kit according to the present embodiment only needs to be able to apply the sample and develop the sample 101 with the developing solution L, has a simple structure, and has a high degree of design freedom.

  The kit according to this embodiment will be described in more detail. The detection target for quantification in this embodiment is 8OHdG, and urine is assumed as a sample including the detection target for quantification. The kit according to the present embodiment is considered to be able to obtain the same effect even with 8-oxo-guanosine, deoxyguanosine, etc. having a guanine skeleton similar to 8OHdG. In this specification, it may be described as 8OHdG, including those that are considered to obtain the same effect.

  It is known that there are various components in urine, especially uric acid is similar in structure to 8OHdG, and the amount of uric acid in urine is several tens to several thousand times that of 8OHdG. ing. Separation from uric acid is necessary to accurately detect and quantify 8OHdG.

  The urine introduced into the introduction unit 102 proceeds to the separation unit 103. The sample is preferably dried once at this stage. The sample of the separation unit 103 includes many components such as 8OHdG and uric acid at this stage. In this embodiment, the method for drying the sample is not particularly limited. However, the sample is allowed to stand for a predetermined period of time to evaporate the moisture naturally, the method of contacting the material that absorbs moisture, or the moisture is evaporated by heating. Methods and the like.

  When a developing solution described later is introduced from the developing solution introduction unit 105 and the developing solution passes through the introducing unit 102, the development of the sample is started. The sample component in the developing solution is repeatedly adsorbed and desorbed with the separation unit 103 which is a solid phase, and is developed toward the detection unit. At this time, by using a developing solution, which will be described later, uric acid as a contaminant is strongly adsorbed by the separation unit 103 that is a solid phase and is not easily developed by the developing solution. On the other hand, 8OHdG is developed together with the developing solution. That is, the development speed of 8OHdG in the development liquid can be made faster than the development speed of impurities such as uric acid and creatinine. Other impurities are developed in different locations from 8OHdG due to the difference in structure with 8OHdG.

  In this way, the sample introduced into the introduction unit 102 is developed and separated at different locations for each component on the separation unit 103.

  Next, the principle that the kit according to the present embodiment quantitatively detects the detection target in the sample will be described with an example. In this example, a case will be described in which the separation unit 103 includes a carrier, and the carrier is silica gel to which an octadecyl group is bonded.

  Since 8OHdG separated on the separation unit 103 is colorless, it cannot be basically confirmed visually. When a fluorescent substance is mixed in advance with the silica gel constituting the separation unit 103, the separation unit 103 is fluorescent by ultraviolet light (UV light). Since many organic compounds containing 8OHdG absorb UV light, the intensity of fluorescence is weak in the region where 8OHdG exists. That is, since the intensity of fluorescence by UV light becomes weaker in the region where the 8OHdG concentration is higher, the 8OHdG concentration can be measured by the detected fluorescence intensity.

  Next, specific examples of each part constituting the test kit according to the present embodiment and materials of each member constituting them will be described.

(sample)
Examples of the sample containing the detection target include urine, blood, sweat, tears, and liquid samples based on these. Moreover, the dilution and concentrated liquid which increased / decreased the quantity of the water to contain are mentioned. Urine is often used as an 8OHdG sample. It is known that the amount of urine 8OHdG is useful as a marker of oxidative stress. Urine contains uric acid, creatinine, urea, various salts, etc. in addition to 8OHdG.

(Support)
The material of the support 104 is made of glass, plastic, aluminum or the like, and can be appropriately selected depending on the type of sample, the type of developing liquid, and the like. As for the size and shape of the support 104, a square having a length of 3 cm to 15 cm and a width of 1 cm to 5 cm in the direction in which the sample moves is preferable for ease of handling. That is, the support is preferably a palm-sized stick from the viewpoint of ease of use.

(Introduction)
The introduction part 102 is a part into which the sample 101 is introduced. A tubular member for introducing the sample 101 onto the introducing portion 102 may be accompanied, or the sample may be introduced by dropping the sample 101 onto the separating portion 103 with a dropper or the like.

  A smaller size of the introduction portion is preferable. If the sample is collected in small spots, the resolution is improved. Separation here means the separation accuracy of each component, and when the separation is high, other components are not mixed with the separated components.

  The shape of the introduction part may be circular or rectangular. In the case of a circle, the diameter is preferably 5 mm or less, and more preferably 3 mm or less. When the rectangle is not a square, the longitudinal direction is preferably 5 mm or longer in the direction perpendicular to the direction in which the developing liquid flows, and more preferably 10 mm or longer. The width in the short direction is preferably 3 mm or less, and more preferably 1 mm or less. When the introduction part is rectangular, the width of the region to which the sample is added is easily narrowed in the direction of progress of the developing solution, and thus the difference in the developing speed between the detection target and the contaminant in the sample is easy to see.

  It is preferable to dry the sample introduced in the introduction part on the spot. By removing the moisture in the sample, the reproducibility of the development with the developing solution is improved, and the detection sensitivity is also improved because the sample is concentrated. You may perform the process of concentrating separately at the time of sample introduction.

(Separation part)
The separation unit 103 includes a carrier (stationary phase) used for chromatography. The carrier is composed of, for example, particulate silica gel or cellulose of about several tens to several hundred microns. The carrier may be bound by a cast agent, an organic polymer, or the like.

  Silica gel may be modified with an organic substance on its surface, for example, silica gel having an octadecyl group bonded thereto may be used. Unmodified silica gel has a strong hydrophilic surface, but a hydrophobic surface can be obtained by modifying the octadecyl group. Silica gel with a hydrophobic surface can easily adsorb uric acid in alcohol and can improve the separation ability from 8OHdG. In addition, the support | carrier may be comprised including the some silica gel from which hydrophobicity mutually differs.

  The separation unit 103 may include a substance that emits fluorescence or phosphorescence, or a coloring substance. If a fluorescent material (fluorescent agent) is contained, a part of the organic matter on the separation unit 103 can be visually recognized with UV light. 8OHdG has a cyclic structure of carbon in its structure and absorbs UV light. Therefore, it can be visually recognized as a black shadow by UV light on the separation part containing the fluorescent material.

  The separation unit 103 may include a component that reacts with 8OHdG and emits fluorescence. Moreover, the component which reacts with 8OHdG and colors may be sufficient. These components can be detected by the detection unit 106.

(Detection unit)
The detection unit 106 is a place where the amount or concentration of 8OHdG is measured. The detection unit 106 needs to visualize the 8OHdG concentration, and examples of the means for visualization include the following.

  (Visualization means 1) The separation unit 103 contains a fluorescent material that absorbs UV light and emits fluorescence, and irradiates the UV light. The higher the 8OHdG concentration, the more visible it is as a black shadow. In this case, the detection illumination 107 is a UV light source.

  (Visualization means 2) After the development, a reagent that reacts with 8OHdG and colors is sprayed on the separation unit 103. Although the color development changes depending on the sprayed reagent, the color development basically follows the 8OHdG concentration. In this case, the detection illumination 107 may not be provided.

  (Visualization means 3) After development, a reagent that reacts with 8OHdG and fluoresces is dispersed in the separation unit 103. Intensity fluorescence according to the 8OHdG concentration is obtained by UV light. In this case, the detection illumination 107 is a UV light source.

(Signal receiver)
Examples of the signal receiving unit 108 for measuring the amount of 8OHdG include the following means.

  (Measuring means 1) The 8OHdG density distribution is analyzed as an image by a digital camera or the like having a CCD or CMOS sensor.

  (Measurement means 2) The amount of reflected light and the amount of fluorescence are measured with a photodiode or the like.

  (Measurement means 3) Visually determine the 8OHdG concentration. Here, an example in which the visualization means 1 and the measurement means 1 are used will be specifically described.

(Quantification of 8OHdG at the detector)
A UV light source of 254 nm is prepared at the upper part of the detection unit, UV light is irradiated, and an 8OHdG shadow is photographed using a digital camera.

When a shadow of 8OHdG is photographed by irradiating UV light, the shadow becomes darker as the amount of 8OHdG increases. FIG. 3 shows an example of a photographed image, where 201 is an introduction unit, 202 is a separation unit, and 203 is a detection unit. In 205, an 8OHdG aqueous solution (for example, 2 μl of 1 mg / ml) having a known concentration is dropped in advance and dried. 203 shows a shadow of 8OHdG in the sample after the sample is introduced into 201 and expanded. 204 is an area not including 8OHdG, and no shadow is shown. First, the R, G, and B values of each pixel in the area 204 are averaged to obtain the average R, average G, and average B values. Next, the average R, average G, and average B values are subtracted from the RGB values of each pixel of the entire image to obtain a difference. At this stage, the R, G, and B values of the pixels are almost zero in the region not including 8OHdG, and have a negative value in the region including 8OHdG. Further, the RGB values of the pixels in the region 203 are integrated to calculate an average value of RGB ((R ^ 2 + G ^ 2 + B ^ 2) ^ (1/2)) (hereinafter, this RGB average value is expressed as sample luminance). ). Similarly, the RGB average value of the pixels in the area 205 is calculated (hereinafter, this RGB average value is referred to as reference luminance). Dividing the sample luminance value by the reference luminance value and multiplying the amount of 8OHdG given to 205 gives information on the concentration of 8OHdG in the sample.

(Holding part, developing liquid introduction part)
The kit in the present embodiment may be configured such that, for example, when a pressure is applied by pressing the holding unit, the developing liquid held in the holding unit flows into the developing liquid introduction unit and flows to the introduction unit. it can.

<Developing solution>
The developing solution according to the present embodiment is a mobile phase in the chromatography and plays a role of developing the detection target in the sample. The developing solution according to the present embodiment is selected such that the developing speed of 8OHdG by the developing solution is faster than the developing speed of impurities.

  For example, if the developing solution is selected such that the solubility of 8OHdG in the developing solution is higher than the solubility of impurities, the relationship between the developing speeds as described above can be realized. Examples of such a developing solution include a solution containing at least one of alcohol and ether and an inorganic electrolyte.

(Main component of developing solution)
In the present embodiment, the main components of the developing solution are alcohol and ether. Examples of the alcohol include at least one of methanol, ethanol, propanol, and isopropanol. Examples of the ether are at least one of dimethyl ether, ethyl methyl ether, diethyl ether, diethylene glycol monoethyl ether, diethylene glycol monomethyl ether, and triethylene glycol monobutyl ether.

  These alcohols have low solubility in substances other than 8OHdG in urine (hereinafter referred to as impurities), particularly uric acid. On the other hand, 8OHdG is optimal for separation because it dissolves.

(Inorganic electrolyte)
Until now, it was thought that even if an inorganic electrolyte was contained in the developing solution, many inorganic electrolytes were adsorbed on the silica gel in the solid phase and did not affect the developing result. Therefore, generally, an inorganic electrolyte has not been used in a developing solution for thin layer chromatography.

  However, as a result of studies by the present inventors, it has been found that the inorganic electrolyte has an effect of suppressing the development of uric acid, particularly uric acid in chromatography. Although the detailed mechanism is unknown, it is thought that the moisture remaining in the main component of the developing solution is adsorbed on the silica gel in the solid phase while the inorganic electrolyte is retained, thereby suppressing the involvement of moisture in uric acid. It is done. It is also considered that uric acid is ionized by the inorganic electrolyte and is easily adsorbed on the silica gel in the solid phase. This effect facilitates separation of 8OHdG and uric acid. The inorganic electrolyte is preferably deliquescent that easily retains moisture. Note that deliquescent means that when the substance is placed in the air, it takes in moisture from the air to form an aqueous solution.

  Examples of the inorganic electrolyte include potassium hydroxide and sodium hydroxide. The concentration of the inorganic electrolyte is preferably 0.09 mol / l or more.

(moisture)
When the developing solution contains water, uric acid is dissolved, so that the separation ability is lowered. Therefore, the water content in the main component of the developing solution is preferably 1.5% or less with respect to the weight of the developing solution. More preferably, it is 1% or less, and more preferably 0.5% or less. Moreover, it is preferable that the molar concentration of water is 3.2 times or less the molar concentration of the inorganic electrolyte. If it is below this concentration, 8OHdG and uric acid can be separated even if water is contained.

(Thin layer chromatography)
The thin layer chromatography for detecting 8 oxo 2 ′ deoxyguanosine according to this embodiment includes the following steps.
(1) A step of introducing a sample containing 8 oxo 2 ′ deoxyguanosine and impurities.
(2) A step of separating and extracting 8OHdG from the sample by developing a developing solution on the sample.
(3) A step of detecting the amount or concentration of the separated and extracted 8 oxo 2 ′ deoxyguanosine.
In addition, before developing a developing liquid, the process of removing the water | moisture content contained in a sample may be included. By removing the water, it becomes easier to separate 8OHdG and uric acid as described above. Moreover, you may further have the process of concentrating a sample. When the sample is urine, the contaminants include at least one of creatinine and uric acid.

(Sample preparation)
Each material was mixed so that it might become content of the following Table 1, 1N sodium hydroxide solution was contained, and it adjusted to 1 ml. Each sample was neutralized with an equal volume of 1N sulfuric acid solution immediately before use.

(Preparation of developing solution)
A developing solution was prepared by containing water and potassium hydroxide in absolute ethanol. Anhydrous ethanol containing molecular sieve 3A was stirred and allowed to stand for 24 hours to sufficiently remove the contained water. The contents of water and potassium hydroxide are as shown in Table 2 below. In either case, the unit is mol / l.

Example 1
A silica gel plate (PLC silica gel RP-18F254 manufactured by Merck Ltd.) cut into 2 cm × 5 cm was used as a separation part. In this silica gel plate, the support is glass, and the silica gel layer in the separation part contains a fluorescent material (fluorescent agent) that absorbs UV. One side of the short side of the silica gel plate was used as the base, and a position 10 mm from the base was used as the introduction part, and 2 μl of sample 1 was applied using a glass tube dropper. The sample was dried on a hot plate at 60 ° C. The developing solution 1 was added to the bottom of the TLC developing layer (manufactured by ASONE) to a depth of about 2 mm, and the developing layer was covered and allowed to stand for 10 minutes. Thereafter, the silica gel plate was placed so that the bottom of the silica gel plate was in the developing layer, and allowed to stand until the developing solution rose 20 mm above the introduction portion. The silica gel plate was taken out, and 2 μl of sample 1 was applied with a glass tube dropper to a position 5 mm from another short side paired with the bottom side (reference). A range of 10 mm from the portion where the sample 1 developing solution was raised toward the bottom was used as a detection unit, and a 254 nm UV light source (handy UV lamp manufactured by ASONE Co., Ltd.) was irradiated and photographed with a digital camera. The photographed image was digitized according to the method of 8OHdG quantification at the detection unit.

(Example 2)
The same experiment as in Example 1 was performed except that the developing solution 2 was used.

(Example 3)
The same experiment as in Example 1 was performed except that the developing solution 3 was used.

  For Samples 2 to 7, the amount of 8OHdG was quantified in the same manner as Sample 1. All references were sample 1. The 8OHdG amount contained at the time of sample preparation was plotted on the horizontal axis in FIG. 3, and the 8OHdG amount determined in the vertical axis was plotted. Samples 1 to 7 were plotted. The amount almost equivalent to 8OHdG contained at the time of preparation was quantified, and 8OHdG could be quantified easily by this method.

(Example 4)
The same experiment as in Example 1 was performed except that the developing solution 4 was used.

(Example 5)
The same experiment as in Example 1 was performed except that the developing solution 5 was used.

(Comparative Example 1)
The same experiment as in Example 1 was performed except that the developing solution 6 was used.

(Evaluation)
When the amount of 8OHdG detected in each sample and the amount of 8OHdG charged at the time of sample preparation are within 10% in all samples, A (preferred level), and in the case where there are many samples with differences exceeding 10%, C (Unacceptable level), otherwise B. The evaluation results are shown in Table 3.

  It was found that 8OHdG could not be detected accurately when potassium hydroxide was not included as in the developing solution 6. Moreover, the quantitative detection of 8OHdG could be accurately performed by setting the potassium hydroxide in the developing solution to an appropriate range.

DESCRIPTION OF SYMBOLS 101 Sample 102 Introduction part 103 Separation part 104 Support body 105 Developing liquid introduction part 106 Detection part 107 Illumination for detection 108 Signal reception part 109 Detection object (8OHdG)
110 Holding part

Claims (28)

An introduction part into which a sample containing 8 oxo-2′deoxyguanosine and impurities is introduced;
A developing solution for developing the 8-oxo-2′-deoxyguanosine and the contaminants of the sample;
A kit for thin-layer chromatography, comprising: a detection unit for detecting the amount or concentration of the 8 oxo-2′deoxyguanosine contained in a solution obtained by developing the sample with the developing solution,
The kit characterized in that the development rate of the 8 oxo-2'deoxyguanosine by the developing solution is faster than the developing rate of the contaminants by the developing solution.   The kit according to claim 1, wherein the solubility of the 8oxo-2'deoxyguanosine in the developing solution is higher than the solubility of the contaminant in the developing solution.   The kit according to claim 1 or 2, wherein the contaminant contains at least one of uric acid and creatinine.   The kit according to any one of claims 1 to 3, wherein the developing solution contains at least one of alcohol and ether and an inorganic electrolyte.   The kit according to any one of claims 1 to 4, wherein the developing solution has a deliquescent inorganic electrolyte.   The kit according to any one of claims 1 to 5, wherein the developing solution contains at least one of potassium hydroxide and sodium hydroxide.   The kit according to any one of claims 1 to 6, wherein the developing solution contains at least one of methanol, ethanol, propanol, and isopropanol. An introduction part into which a sample containing a detection object is introduced;
A developing solution for developing the detection object of the sample;
A detection unit for detecting the amount or concentration of the detection target contained in a solution obtained by developing the sample with the developing solution, and a kit for thin layer chromatography,
The developing solution contains at least one of alcohol and ether and at least one of potassium hydroxide and sodium hydroxide. The alcohol includes at least one of methanol, ethanol, propanol, and isopropanol, and the ether is at least one of dimethyl ether, ethyl methyl ether, diethyl ether, diethylene glycol monoethyl ether, diethylene glycol monomethyl ether, and triethylene glycol monobutyl ether. The kit according to claim 8, comprising at least one of them.   The kit according to claim 9, wherein the detection target is 8 oxo-2'deoxyguanosine.   The kit according to any one of claims 1 to 10, further comprising a holding unit that holds the developing solution.   The kit according to any one of claims 1 to 7, wherein a carrier for adsorbing the contaminants is provided between the introduction unit and the detection unit.   The kit according to any one of claims 8 to 10, wherein a carrier is provided between the introduction part and the detection part.   The kit according to claim 12 or 13, wherein the carrier comprises at least one of silica gel and cellulose.   The kit according to claim 12 or 13, wherein the carrier comprises silica having an octadecyl group bonded thereto.   The kit according to claim 12 or 13, wherein the carrier comprises a plurality of silica gels having different hydrophobicity from each other.   The kit according to any one of claims 1 to 16, wherein the detection unit is provided with a fluorescent material that absorbs ultraviolet light.   The said detection part is provided with the substance which reacts with the said 8 oxo 2 'deoxy guanosine, and emits fluorescence or phosphorescence, or a color development substance, It is any one of Claim 1 thru | or 7 characterized by the above-mentioned. kit.   The kit according to any one of claims 8 to 10, wherein the detection unit is provided with a substance that emits fluorescence or phosphorescence by reacting with the detection object, or a coloring substance.   A developing solution for thin layer chromatography, comprising at least one of alcohol and ether and an inorganic electrolyte.   The developing solution for thin layer chromatography according to claim 20, wherein the inorganic electrolyte is deliquescent.   The developing solution for thin layer chromatography according to claim 20 or 21, wherein the alcohol is at least one of methanol, ethanol, propanol and isopropanol.   The developing solution for thin layer chromatography according to any one of claims 20 to 22, wherein the inorganic electrolyte is at least one of potassium hydroxide and sodium hydroxide.   The molar concentration of water contained in the developing solution is not more than 3.2 times the molar concentration of the inorganic electrolyte, The thin layer chromatography according to any one of claims 20 to 23, Developing liquid.   The developing solution for thin-layer chromatography according to any one of claims 20 to 24, wherein the concentration of the inorganic electrolyte contained in the developing solution is 0.09 mol / l or more.   Thin-layer chromatography for detecting 8oxo-2′deoxyguanosine, the step of introducing a sample containing the 8oxo2′deoxyguanosine and impurities, and developing a developing solution on the sample, A thin-layer chromatography comprising: a step of separating and extracting 8OHdG; and a step of detecting the amount or concentration of the 8oxo-2′deoxyguanosine separated and extracted.   27. The thin layer chromatography according to claim 26, wherein the contaminant contains at least one of creatinine and uric acid.   The thin layer chromatography according to claim 26 or 27, further comprising a step of concentrating the sample.

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