JPH0145028B2 - - Google Patents

Info

Publication number
JPH0145028B2
JPH0145028B2 JP20349182A JP20349182A JPH0145028B2 JP H0145028 B2 JPH0145028 B2 JP H0145028B2 JP 20349182 A JP20349182 A JP 20349182A JP 20349182 A JP20349182 A JP 20349182A JP H0145028 B2 JPH0145028 B2 JP H0145028B2
Authority
JP
Japan
Prior art keywords
weight
latex
latex particles
particles
penicillamine
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired
Application number
JP20349182A
Other languages
Japanese (ja)
Other versions
JPS5992354A (en
Inventor
Michio Kuge
Micha Nakagawa
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Sekisui Chemical Co Ltd
Original Assignee
Sekisui Chemical Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Sekisui Chemical Co Ltd filed Critical Sekisui Chemical Co Ltd
Priority to JP20349182A priority Critical patent/JPS5992354A/en
Publication of JPS5992354A publication Critical patent/JPS5992354A/en
Publication of JPH0145028B2 publication Critical patent/JPH0145028B2/ja
Granted legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form

Landscapes

  • Health & Medical Sciences (AREA)
  • Immunology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Hematology (AREA)
  • Urology & Nephrology (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Cell Biology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Description

【発明の詳細な説明】[Detailed description of the invention]

本発明は免疫血清学的診断試薬に関する。 従来、ラテツクス粒子を担体とし、抗原又は抗
体を感作させ、血清中の抗体もしくは抗原と特異
的に起る抗原、抗体反応によりラテツクス粒子の
凝集反応を生じさせ、その結果により各種疾患の
診断を行うことが免疫血清学的診断法として臨床
検査の分野において行なわれており、例えばリウ
マチ因子、HBs抗原、HBs抗体、抗ストレプト
リジン−Ｏ（ASO）、Ｃ−反応性蛋白質（CRP）、
α−フエトプロテイン、癌胎児性抗原（CEA）
等の検査にもこのような免疫血清学的診断試薬が
用いられている。 かゝる診断試薬として、例えばポリスチレン粒
子が分散されているラテツクス懸濁液中の、ポリ
スチレン粒子にガンマグロブリンを混合し粒子表
面に吸着させたものが一般に使用されている。こ
のような診断試薬では、ガンマグロブリンは溶液
中でラテツクス粒子に吸着されずに存在するもの
とラテツクス表面上のものとが吸脱着して平衡状
態にある。そしてガンマグロブリンは主体がイム
ノグロブリンＧ（IgG）であり、ガンマグロブリ
ン中の90％以上を占めている。IgGは分子同志が
会合し２量体、３量体等の多量体を形成しやすい
が、かゝる多量体はラテツクス粒子を凝集させる
性質が強く、試薬の経時変化をまねくおそれがあ
る。 このため例えば特開昭54−26327号公報ではラ
テツクス懸濁液中に塩化コリン、サツカロースを
加えることにより経時変化を抑制している。しか
し塩化コリン、サツカロースを添加すると懸濁液
の粘度上昇をきたしラテツクス粒子の凝集反応に
際しての感度の低下、すなわち検出可能な測定対
象物の濃度下限の上昇をきたし、又凝集反応速度
が低下する欠点が新たに生ずる。 本発明はこのような欠点を解消し、ラテツクス
粒子の凝集反応に際しての感度が長期間に亘り高
感度に維持され、長期間の保存によつても自己凝
集が生じない免疫血清学的診断試薬を提供するこ
とを目的とする。 本発明の要旨は、粒径0.08μｍから0.80μｍのラ
テツクス粒子10mg当り、ガンマグロブリンが
250μｇから5000μｇ感作されており、該ラテツク
ス粒子が0.08重量％から2.0重量％含有されてい
るラテツクス懸濁液からなり、該懸濁液中に0.01
重量％から10.0重量％のペニシラミンが添加され
ていることを特徴とする、免疫血清学的診断試薬
に存する。 次に本発明免疫血清学的診断試薬について更に
詳細に説明する。 本発明におけるラテツクス粒子の粒径は、
0.08μｍから0.80μｍの範囲にあるものが使用され
る。これは、ラテツクス粒子に感作されたガンマ
グロブリンが検体中の抗原又は抗体と反応して凝
集反応を生ずるとラテツクス粒子が数個乃至数十
個くつついて凝集像を生ずるが、これをガラス板
上で肉眼判定したり、光学的に散乱光の変化でと
らえたり、透過光の強弱変化でとらえたりするに
適当な粒径範囲である。 ラテツクス粒子は、全粒子が0.08μｍから0.80μ
ｍの範囲に存在するものが好ましいが、凝集像に
影響を与えない程度の少量、望ましくは５％以内
でかゝる粒径から若干外れるものを含むことも許
容される。 ラテツクス粒子としては、例えばポリスチレン
粒子の他、フエニル基を有する単量体と陰イオン
性単量体との共重合体粒子が好適である。 共重合体粒子の場合、フエニル基を有する単量
体としては、例えばスチレン、ジビニルベンゼ
ン、エチルスチレン、α−メチルスチレン等が存
し、又陰イオン性の単量体としては、例えばスチ
レンスルホン酸塩、ジビニルベンゼンスルホン酸
塩、エチルスチレンスルホン酸塩、α−メチルス
ルホン酸塩等が存する。 又、前記の共重合体粒子を得るには、例えばフ
エニル基を有する単量体100重量部と陰イオン性
単量体0.1〜70重量部を孔化剤の不存在下に過硫
酸塩を重合開始剤として水中で共重合反応を生じ
させることにより得られる。このようにして得ら
れる共重合体粒子には、共重合成分である陰イオ
ン性単量体に基づくものと重合開始剤の切片の陰
イオンに基づく表面荷電が付与される。共重合体
粒子の表面荷電密度はラテツクス懸濁液における
陰イオンの解離濃度で3.0×10-7モル／m2〜80.0×
10-7モル／m2の範囲内に存するものとされ、かゝ
る範囲内にある場合は、孔化剤が存在しなくとも
共重合体粒子間の自己凝集を防ぎ安定なラテツク
ス懸濁液が得られる。 本発明においてはラテツクス粒子10mg当り、ガ
ンマグロブリンが250μｇから5000μｇ感作され
る。 本発明においてラテツクス粒子が感作されるガ
ンマグロブリンは、ガンマグロブリンそのもので
あつもよいし、又IgG分画であつてもよいし、又
更にはIgGを更に分画して得られる抗原又は抗体
そのものであつてもよい。 例えばCRPを検出する場合において、IgGを更
に分画したＦ（ab′）2が使用されることによりRA
因子の干渉や補体の妨害を除去することができ
る。この場合はＦ（ab′）2の感作量が上記範囲内に
存在するものとされる。 ガンマグロブリンの感作がラテツクス粒子10mg
当り250μｇから5000μｇとされるのは、250μｇよ
りも少量では凝集速度が遅くなるし、凝集反応を
行つた後の凝集像も検出し難くなり、又5000μｇ
より多量では系中に遊離して存在するガンマグロ
ブリンが多く反応するため、結局凝集反応におけ
る感度低下を来たしやすいからである。 ラテツクス懸濁液中に分散されているラテツク
ス粒子の量は0.08重量％から2.0重量％とされる。
ラテツクス粒子の量が上記範囲内に存するように
なされるのは、ラテツクス粒子の量が0.08重量％
よりも少量では凝集反応を行なつた後の凝集像の
検出が困難となり、又2.0重量％よりも多量では
凝集反応により小凝集が多数生じこれがために凝
集像の判定が困難になるからである。 本発明にはラテツクス懸濁液中に0.01重量％か
ら10.0重量％のペニシラミンが添加されている。
ペニシラミンは、次式 で表わされる含硫α−アミノ酸である。 そしてペニシラミンは、IgGの会合を抑制し、
ラテツクス診断試薬の感度低下を防ぐ働きを有す
ることがわかつた。ペニシラミンはＤ体、Ｌ体、
DL体のいずれもが使用できる。ペニシラミンの
添加量は、ラテツクス懸濁液中に0.01重量％から
10.0重量％の範囲とされる。これは0.01重量％よ
りも少量ではIgGの会合抑制や診断試薬として使
用する際の感度低下を防ぐ働きが発現されないお
それがあり、10.0重量％以上含有する場合は薬と
して実際に使用する際試薬表面や試薬の拡がりの
端部が乾燥しやすいがこの場合に溶解度との関係
で結晶を生じやすいものとなるので好ましくない
ことによる。 本発明において診断試薬の感度、凝集反応時間
等を調整するために、ペニシラミンの他に、例え
ば塩化コリン、サツカロース等を添加することも
ありうる。 本発明によれば、ラテツクス粒子の凝集反応に
際しての感度が長期間に亘り高感度に保持される
と共に、長時間の保存によつても自己凝集を生じ
ない免疫血清学的診断試薬が得られる。 実施例 １ (1) Ｂ型肝炎ウイルス表面抗原検出用ラテツクス
試薬の調整、0.015Mのリン酸食塩緩衝液（PH
7.4）中に、粒径が0.46μｍのポリスチレンラテ
ツクス粒子が2.0重量％含有されるように分散
させたもの（Ａ液）を調製した。HBs抗原を
モルモツトに免疫して得られた抗血清から精製
した、抗HBs抗原抗体を含むIgG分画成分を、
0.015Mのリン酸食塩緩衝液（PH7.4）に対して
500μｇ／mlとなるように溶解させたもの（Ｂ
液）を調整した。 このＡ液、Ｂ液を体積比で１：１に混合し、
ポリスチレンラテツクス粒子10mg当り、前記
IgG分画を250μｇ感作し37℃で120分間加熱し
た。 このラテツクス懸濁液に更にペニシラミン、
防腐剤としてアジ化ナトリウムを加え、最終的
にポリスチレンラテツクス粒子が1.0重量％、
ペニシラミン0.5重量％、アジ化ナトリウム0.1
重量％を含有する診断試薬を得た。 (2) 感度判定及び保存安定性試験 上記のようにして得られた診断試薬50μ
と、精製HBs抗原を正常人プール血清で希釈
した検体50μとを凝集判定用スライドグラス
上で混合し３分後の凝集像を肉眼で観察し感度
を判定した。又、診断試薬を調整した直後のも
のと４℃で１年間保存したものについて感度を
比較し保存安定性を評価した。 The present invention relates to immunoserological diagnostic reagents. Conventionally, latex particles have been used as carriers to sensitize antigens or antibodies, and the agglutination reaction of the latex particles is caused by the antigen/antibody reaction that occurs specifically with antibodies or antigens in serum, and the results can be used to diagnose various diseases. This method is used in the field of clinical testing as an immunoserological diagnostic method, such as rheumatoid factor, HBs antigen, HBs antibody, anti-streptolysin-O (ASO), C-reactive protein (CRP),
alpha-fetoprotein, carcinoembryonic antigen (CEA)
Such immunoserological diagnostic reagents are also used for such tests. As such a diagnostic reagent, for example, a latex suspension in which polystyrene particles are dispersed, in which gamma globulin is mixed with polystyrene particles and adsorbed onto the particle surface, is generally used. In such a diagnostic reagent, gamma globulin exists in an equilibrium state in which the gamma globulin that is not adsorbed to the latex particles in the solution and the gamma globulin that is on the latex surface are adsorbed and desorbed. Gamma globulin is mainly composed of immunoglobulin G (IgG), which accounts for more than 90% of gamma globulin. The molecules of IgG tend to associate with each other to form multimers such as dimers and trimers, but such multimers have a strong tendency to aggregate latex particles and may cause deterioration of the reagent over time. For this reason, for example, in JP-A-54-26327, changes over time are suppressed by adding choline chloride and sucrose to a latex suspension. However, the addition of choline chloride and sutucarose increases the viscosity of the suspension, resulting in a decrease in sensitivity during the aggregation reaction of latex particles, i.e., an increase in the lower limit of the concentration of the detectable analyte, and a decrease in the rate of agglutination reaction. is newly created. The present invention eliminates these drawbacks and provides an immunoserological diagnostic reagent that maintains high sensitivity during the agglutination reaction of latex particles over a long period of time and does not cause self-aggregation even after long-term storage. The purpose is to provide. The gist of the present invention is that gamma globulin is
It consists of a latex suspension containing 250 μg to 5000 μg of sensitized latex particles and 0.08% to 2.0% by weight of the latex particles.
An immunoserological diagnostic reagent characterized in that penicillamine is added in an amount of 10.0% by weight. Next, the immunoserological diagnostic reagent of the present invention will be explained in more detail. The particle size of the latex particles in the present invention is
A material in the range of 0.08 μm to 0.80 μm is used. This is because when gamma globulin sensitized by latex particles reacts with the antigen or antibody in the sample and causes an agglutination reaction, several to dozens of latex particles are pecked together to form an agglutinated image, which is then placed on a glass plate. This is an appropriate particle size range for determining with the naked eye, optically detecting changes in scattered light, or detecting changes in the intensity of transmitted light. Latex particles have a total particle size of 0.08μm to 0.80μm.
It is preferable that the particles exist within the range of m, but it is also permissible to include particles slightly deviating from the above range of particle size, preferably within 5%, so as not to affect the agglomerated image. Suitable latex particles include, for example, polystyrene particles and copolymer particles of a monomer having a phenyl group and an anionic monomer. In the case of copolymer particles, examples of monomers having phenyl groups include styrene, divinylbenzene, ethylstyrene, α-methylstyrene, etc., and examples of anionic monomers include styrene sulfonic acid. There are salts, divinylbenzenesulfonate, ethylstyrenesulfonate, α-methylsulfonate, and the like. In addition, in order to obtain the above copolymer particles, for example, 100 parts by weight of a monomer having a phenyl group and 0.1 to 70 parts by weight of an anionic monomer are polymerized with persulfate in the absence of a pore-forming agent. It is obtained by causing a copolymerization reaction in water as an initiator. The copolymer particles thus obtained are given surface charges based on the anionic monomer as a copolymerization component and the anion of the polymerization initiator fragment. The surface charge density of copolymer particles is 3.0×10 -7 mol/m 2 to 80.0× at the dissociated concentration of anions in the latex suspension.
10 -7 mol/m 2 , and if it is within such a range, self-aggregation between copolymer particles can be prevented and a stable latex suspension can be obtained even in the absence of a pore-forming agent. is obtained. In the present invention, 250 to 5000 μg of gamma globulin is sensitized per 10 mg of latex particles. The gamma globulin to which the latex particles are sensitized in the present invention may be gamma globulin itself, an IgG fraction, or even an antigen or antibody itself obtained by further fractionating IgG. It may be. For example, when detecting CRP, F(ab') 2 , which is a further fraction of IgG, is used to detect RA.
Factor interference and complement interference can be removed. In this case, the sensitizing amount of F(ab') 2 is assumed to be within the above range. Gamma globulin sensitization with latex particles 10mg
The reason why the amount is set from 250μg to 5000μg is that if the amount is less than 250μg, the aggregation rate will be slow and the agglutination image after the agglutination reaction will be difficult to detect.
This is because if the amount is too large, a large amount of gamma globulin that is free in the system reacts, which tends to result in a decrease in sensitivity in the agglutination reaction. The amount of latex particles dispersed in the latex suspension is between 0.08% and 2.0% by weight.
The amount of latex particles is within the above range so that the amount of latex particles is 0.08% by weight.
If the amount is less than 2.0% by weight, it will be difficult to detect the agglutination image after the agglutination reaction, and if the amount is more than 2.0% by weight, the agglutination reaction will cause many small aggregations, which will make it difficult to judge the agglutination image. . In the present invention, 0.01% to 10.0% by weight of penicillamine is added to the latex suspension.
Penicillamine has the following formula: It is a sulfur-containing α-amino acid represented by And penicillamine suppresses the association of IgG,
It was found that it has the ability to prevent the sensitivity of latex diagnostic reagents from decreasing. Penicillamine is D-form, L-form,
Any DL version can be used. The amount of penicillamine added starts from 0.01% by weight in the latex suspension.
The range is 10.0% by weight. If the amount is less than 0.01% by weight, there is a risk that the function of suppressing the association of IgG and preventing a decrease in sensitivity when used as a diagnostic reagent will not be expressed.If it is contained more than 10.0% by weight, the surface of the reagent will This is because the ends of the spread of the reagent tend to dry out easily, which is not preferable because in this case crystals tend to form due to the relationship with solubility. In the present invention, in order to adjust the sensitivity, agglutination reaction time, etc. of the diagnostic reagent, in addition to penicillamine, for example, choline chloride, sutucarose, etc. may be added. According to the present invention, it is possible to obtain an immunoserological diagnostic reagent that maintains high sensitivity during the agglutination reaction of latex particles over a long period of time, and does not cause self-aggregation even after long-term storage. Example 1 (1) Preparation of latex reagent for detection of hepatitis B virus surface antigen, 0.015M phosphate saline buffer (PH
7.4), polystyrene latex particles having a particle size of 0.46 μm were dispersed to contain 2.0% by weight (Liquid A). The IgG fraction containing anti-HBs antigen antibodies was purified from the antiserum obtained by immunizing guinea pigs with HBs antigen.
For 0.015M phosphate saline buffer (PH7.4)
Dissolved to a concentration of 500 μg/ml (B
liquid) was adjusted. Mix these A liquid and B liquid at a volume ratio of 1:1,
per 10 mg of polystyrene latex particles,
250 μg of the IgG fraction was sensitized and heated at 37° C. for 120 minutes. In addition, penicillamine is added to this latex suspension.
Sodium azide was added as a preservative, and the final polystyrene latex particles were 1.0% by weight.
Penicillamine 0.5% by weight, sodium azide 0.1
A diagnostic reagent containing % by weight was obtained. (2) Sensitivity determination and storage stability test 50μ of the diagnostic reagent obtained as above
and 50μ of a sample prepared by diluting purified HBs antigen with normal human pool serum were mixed on a slide glass for agglutination determination, and the agglutination image after 3 minutes was observed with the naked eye to determine sensitivity. In addition, the storage stability was evaluated by comparing the sensitivity of the diagnostic reagents immediately after preparing them and those stored at 4° C. for one year.

【表】【table】

【表】 第１表から明らかなように、診断試薬を調整
直後のものと、調整後４℃で１年保存後のもの
とでは凝集像の感度に変化がなく、保存安定性
がすぐれていることが確認された。 比較例 １ 実施例１との比較の為に、実施例１においてペ
ニシラミンが使用されないものについて同様に試
験を行つた結果では４℃で６ケ月後経過後に精製
HBs抗原濃度が10- 2μｇ／mlの場合に＋に判定さ
れた。 この場合のラテツクス試薬は陰性コントロール
（HBs抗原を含まない正常人血清）に対しても＋
に判定されるおそれがあり、診断試薬として使用
できないものであつた。 実施例 ２ (1) リウマチ因子検出用ラテツクス試薬の調整 粒径が0.20μｍのポリスチレンラテツクス粒
子をグリシン−カセイソーダ緩衝液に分散させ
たラテツクス懸濁液における、ポリスチレンラ
テツクス粒子10mg当り3000μｇのヒトガンマグ
ロブリンを感作し４℃で１夜放置した。このラ
テツクス懸濁液にペニシラミン、牛血清アルブ
ミン、を加え、更に防腐剤としてアジ化ナトリ
ウムを加え、最終的にポリスチレンラテツクス
粒子0.5重量％、ペニシラミン10.0重量％、牛
血清アルブミン1.0重量％、アジ化ナトリウム
0.1重量％が夫々含有されている診断試薬を得
た。 (2) 感度判定及び保存安定性試験 10個の血清検体について実施例１におけると
同様にして感度判定及び保存安定性試験を行つ
た結果を第２表に示す。[Table] As is clear from Table 1, there is no change in the sensitivity of the agglutination image between the diagnostic reagent immediately after preparation and the one after storage at 4℃ for 1 year, and the storage stability is excellent. This was confirmed. Comparative Example 1 For comparison with Example 1, a test was conducted in the same manner as in Example 1 in which penicillamine was not used, and the results showed that the product was purified after 6 months at 4°C.
It was judged as positive when the HBs antigen concentration was 10 - 2 μg/ml. In this case, the latex reagent is also positive against the negative control (normal human serum that does not contain HBs antigen).
Therefore, it could not be used as a diagnostic reagent. Example 2 (1) Preparation of latex reagent for detecting rheumatoid factor 3000 μg of human gamma per 10 mg of polystyrene latex particles in a latex suspension in which polystyrene latex particles with a particle size of 0.20 μm were dispersed in a glycine-caustic soda buffer. The cells were sensitized with globulin and left overnight at 4°C. Penicillamine, bovine serum albumin, and sodium azide are added to this latex suspension, and sodium azide is added as a preservative.Finally, 0.5% by weight of polystyrene latex particles, 10.0% by weight of penicillamine, 1.0% by weight of bovine serum albumin, and azide are added. sodium
Diagnostic reagents containing 0.1% by weight of each were obtained. (2) Sensitivity determination and storage stability test Sensitivity determination and storage stability test were performed on 10 serum samples in the same manner as in Example 1. The results are shown in Table 2.

【表】 第２表から明らかなように、診断試薬を調整
直後のものと調整後４℃で１年間保存後のもの
とでは凝集像の感度に変化がなく、保存安定性
がすぐれていることが確認された。 実施例 ３ (1) CRP検出用ラテツクス試薬の調整 粒径が0.12μｍのポリスチレンラテツクス粒
子をグリシン−カセイソーダ緩衝液（PH8.6）
に分散させたラテツクス懸濁液における、ポリ
スチレンラテツクス粒子10mg当り、ヤギ産生の
抗ヒトCRP抗血清から精製分離したIgG分画
500μｇを感作した。 次いでこのラテツクス懸濁液にペニシラミ
ン、サツカロース、牛血清アルブミン、塩化コ
リン、アジ化ナトリウムを夫々加え、最終的に
ポリスチレンラテツクス粒子0.8重量％、ペニ
シラミン0.01重量％、サツカロース1.0重量％、
牛血清アルブミン1.0重量％、塩化コリン2.5重
量％、アジ化ナトリウム0.1重量％が夫々含有
されている診断試薬を得た。 (2) 感度判定及び保存安定性試験 上記のようにして得られた診断試薬につい
て、精製CRPの含有濃度の異なる血清検体に
ついて、実施例１と同様にして感度判定及び保
存安定性試験を行つた結果、第３表に示すよう
に保存安定性にすぐれていることが確認され
た。[Table] As is clear from Table 2, there is no change in the sensitivity of the agglutination image between the diagnostic reagents immediately after preparation and those stored at 4°C for one year after preparation, indicating excellent storage stability. was confirmed. Example 3 (1) Preparation of latex reagent for CRP detection Polystyrene latex particles with a particle size of 0.12 μm were mixed with glycine-caustic soda buffer (PH8.6)
IgG fraction purified and separated from goat-produced anti-human CRP antiserum per 10 mg of polystyrene latex particles in a latex suspension dispersed in
500 μg was sensitized. Next, penicillamine, sutucarose, bovine serum albumin, choline chloride, and sodium azide were added to this latex suspension, and finally polystyrene latex particles 0.8% by weight, penicillamine 0.01% by weight, sutucarose 1.0% by weight,
A diagnostic reagent containing 1.0% by weight of bovine serum albumin, 2.5% by weight of choline chloride, and 0.1% by weight of sodium azide was obtained. (2) Sensitivity determination and storage stability test Regarding the diagnostic reagent obtained as described above, sensitivity determination and storage stability test were conducted in the same manner as in Example 1 using serum samples containing different concentrations of purified CRP. As a result, as shown in Table 3, it was confirmed that the product had excellent storage stability.

【表】 比較例 ３ 実施例３との比較の為に、実施例３においてペ
ニシラミンが除かれている試薬を調整し、実施例
３と同様に感度判定及び保存安定性試験を行つた
結果第４表に示すように保存安定性の低下を来た
すことが認められた。[Table] Comparative Example 3 For comparison with Example 3, a reagent was prepared in which penicillamine was removed from Example 3, and the sensitivity determination and storage stability test were conducted in the same manner as in Example 3. As shown in the table, it was observed that storage stability decreased.

【表】 実施例 ４ (1) CRP検出用ラテツクス試薬の調整 粒径が0.12μのスチレン−スチレンスルホン
酸ソーダ共重合体ラテツクス粒子が5.0重量％
（50mg／ml）含有されるように、前記共重合体
ラテツクス粒子をリン酸食塩緩衝液（PH6.5）
に分散させたもの（Ａ液）を調整した。 ヤギ産生の抗ヒトCRP抗血清から精製分離
したIgG分画をリン酸食塩緩衝液（PH6.5）に
5000μｇ／ml溶解したもの（Ｂ液）を調整し
た。 前記Ａ液、Ｂ液を体積比で１：１に混合し、
37℃で180分間撹拌し、前記共重合体ラテツク
ス粒子10mg当りIgG分画1000μｇを感作させた。
更にペニシラミン、アジ化ナトリウムをリン酸
食塩緩衝液（PH6.5）の溶液として加え、最終
的に前記共重合体ラテツクス粒子が1.0重量％、
ペニシラミン5.0重量％、アジ化ナトリウム0.1
重量％を含有する診断試薬を得た。尚ラテツク
ス粒子の表面荷電密度はラテツクス懸濁液にお
ける陰イオンの解離濃度で16.4×10-7モル／m2
であつた。 (2) 感度判定及び保存安定性試験 上記のようにして得られた診断試薬につい
て、精製CRPの含有濃度の異なる血清検体に
ついて、実施例１と同様にして感度判定及び保
存安定性試験を行つたところ、第５表に示すよ
うに保存安定性にすぐれていることが確認され
た。[Table] Example 4 (1) Preparation of latex reagent for CRP detection 5.0% by weight of styrene-sodium styrene sulfonate copolymer latex particles with a particle size of 0.12μ
(50 mg/ml) of the copolymer latex particles in phosphate saline buffer (PH6.5).
(Liquid A) was prepared. IgG fraction purified from goat-produced anti-human CRP antiserum was added to phosphate saline buffer (PH6.5).
A solution containing 5000 μg/ml (solution B) was prepared. Mixing the A liquid and B liquid at a volume ratio of 1:1,
The mixture was stirred at 37° C. for 180 minutes to sensitize 1000 μg of the IgG fraction per 10 mg of the copolymer latex particles.
Furthermore, penicillamine and sodium azide were added as a solution in phosphate saline buffer (PH6.5), and finally the copolymer latex particles were 1.0% by weight.
Penicillamine 5.0% by weight, sodium azide 0.1
A diagnostic reagent containing % by weight was obtained. The surface charge density of latex particles is the dissociated concentration of anions in the latex suspension, which is 16.4×10 -7 mol/m 2
It was hot. (2) Sensitivity determination and storage stability test Regarding the diagnostic reagent obtained as described above, sensitivity determination and storage stability test were conducted in the same manner as in Example 1 using serum samples containing different concentrations of purified CRP. However, as shown in Table 5, it was confirmed that the product had excellent storage stability.

【表】【table】

Claims (1)

【特許請求の範囲】[Claims] １ 粒径0.08μｍから0.80μｍのラテツクス粒子10
mg当り、ガンマグロブリンが250μｇから5000μｇ
感作されており、該ラテツクス粒子が0.08重量％
から2.0重量％含有されているラテツクス懸濁液
からなり、該懸濁液中に0.01重量％から10.0重量
％のペニシラミンが添加されていることを特徴と
する免疫血清学的診断試薬。1 Latex particles with a particle size of 0.08 μm to 0.80 μm 10
Gamma globulin is 250μg to 5000μg per mg
Sensitized, the latex particles contain 0.08% by weight
An immunoserological diagnostic reagent comprising a latex suspension containing penicillamine in an amount of 2.0% by weight from 0.01% to 10.0% by weight.
JP20349182A 1982-11-18 1982-11-18 Reagent for immune serological diagnosis Granted JPS5992354A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
JP20349182A JPS5992354A (en) 1982-11-18 1982-11-18 Reagent for immune serological diagnosis

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
JP20349182A JPS5992354A (en) 1982-11-18 1982-11-18 Reagent for immune serological diagnosis

Publications (2)

Publication Number Publication Date
JPS5992354A JPS5992354A (en) 1984-05-28
JPH0145028B2 true JPH0145028B2 (en) 1989-10-02

Family

ID=16475032

Family Applications (1)

Application Number Title Priority Date Filing Date
JP20349182A Granted JPS5992354A (en) 1982-11-18 1982-11-18 Reagent for immune serological diagnosis

Country Status (1)

Country Link
JP (1) JPS5992354A (en)

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS62218864A (en) * 1986-03-20 1987-09-26 Hitachi Chem Co Ltd Quantitative analysis of human c reactive protein
JPS62218866A (en) * 1986-03-20 1987-09-26 Hitachi Chem Co Ltd Reagent for quantitative analysis of human c-reactive protein
JPH0833382B2 (en) * 1986-03-28 1996-03-29 エスエイ・テキサコ・ベルジヤン・エヌヴイ probe
JP2534067B2 (en) * 1987-07-09 1996-09-11 日水製薬株式会社 Quantitative method for C-reactive protein
JP6224217B1 (en) * 2016-12-27 2017-11-01 Jsr株式会社 How to store latex particle dispersion

Also Published As

Publication number Publication date
JPS5992354A (en) 1984-05-28

Similar Documents

Publication Publication Date Title
US4656144A (en) Immunoparticles and process for preparing same
US4060597A (en) Serological reagent and preparation thereof
CN108593641A (en) A kind of kit and method quantitatively detecting test substance in whole blood sample
US4552633A (en) Fine particulate for use in clinical testing and a process for producing thereof
JPH0810224B2 (en) A latex for immobilizing a physiologically active substance and a latex reagent using this latex
CN101354400B (en) Reagent for latex aggregation reaction and method for detecting target substance
JP3471198B2 (en) Reagent for turbidimetric determination of immunological latex
EP2902785B1 (en) Latex particles for particle aggregation measurement
JPS5992354A (en) Reagent for immune serological diagnosis
JPS5846243B2 (en) Method for producing latex for serological diagnostic reagents
JP2545707B2 (en) Immunological diagnostic reagent
JPH02257063A (en) Reagent and method for immunoassay
JPS5850646B2 (en) Method for producing latex for serological diagnostic reagents
JPH0157688B2 (en)
JPS6116944B2 (en)
JP2003344410A (en) Immunoassay reagents and immunoassays
JPS6116942B2 (en)
CA2088471C (en) Method for determining rheumatoid factors and agents for carrying out the method
JPS6116941B2 (en)
JP2545503B2 (en) Immunological diagnostic reagent
JPS6315554B2 (en)
JPS6352706B2 (en)
JPH0564744B2 (en)
JPS5834486B2 (en) Method for producing latex for diagnostic reagents
JPS6352708B2 (en)