マウス肝臓リボソームは1.8%ポリアクリルアミドー0.5%アガロース混成ゲル (7.5% cross linkage gel) 電気泳動で分析された. リボソーム懸濁液をアガロースと混ぜて, ゲル溝に添加して, ゲルの濃度を変えないで, 二次元電気泳動 (turned gel electrophoresis) を行うと, 40Sサブユニットから最大80Sリボソームの六量体までのリボソームが分離されることが認められた. この分解能を高くした電気泳動は非変性のリボヌクレオ蛋白粒子を分析したり, 抗生物質によるリボソームの変化を研究する有力な手段の一つになると考える.

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Several dyes were evaluated for their potential use as indices of ionic boundaries and pH discontinuities during polyacrylamide gel electrophoresis (PAGE). Dye selection was based largely upon color transition over the pH intervals present in the commonly used Tris-HCl-glycine buffer systems. In the absence of sodium dodecylsulfate (SDS) nitrazine yellow, cresol purple, cresol red, and phenol red had relative mobilities (relative to the chloride boundary; R_C1) of 1.00 at 20% polyacrylamide concentration (% T) compared to bromophenol blue (BPB) for which R_C1=0.77. The homologue of BPB, 3, 4, 5, 6-tetrabromophenolsulfonephthalein (TBPS), exhibited a useful color transition over the pH 6.6-8.2 range but was an equally poor indicator of the chloride boundary at 20% T in the absence of SDS (R_C1=0.73). The R_C1 of other dyes was increased in the presence of SDS, particularly the cationic dye neutral red which complexes with micellar but not monomeric SDS. In the presence of SDS, all dyes except neutral red exhibited R_C1 of 1.00 at 15% T, whereas only neutral red and nitrazine yellow had R_C1 values of 1.00 at lower gel concentrations. Partial characterization of SDS, in terms of R_C1 and zone broadening, was performed in a system devoid of extraneous SDS in the gel or electrolytes.

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The behaviors of all the components of glycine and tricine sodium dodecyl sulfate-polyacrylamide gel electrophoresis system in the course of separation of polypeptide-dodecyl sulfate complexes are investigated by computer simulation. In both of the systems, the leading chloride ion zone is followed by concentrated dodecyl sulfate zone, the length of which gradually increases with the time of electrophoresis. Under the postulated conditions and the values of physico-chemical parameters of the components of the systems, almost all the polypeptides migrate as the dodecyl sulfate-polypeptide complexes. These complexes are stacked between the dodecyl sulfate zone and the terminating glycine or tricine zone when they reach the separation gel. The high mobility complexes are not overtaken by the terminating glycine in the separation gel but stacked between the dodecyl sulfate and the glycine zone, while, all the complexes are overtaken in the separation gel by the terminating tricine and separated in the separation gel. Thus, the reason for the usefulness of tricine for the separation of the low molecular complexes is clarified by the computer simulation.

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There are only a few reports concerning analyses of the protein components in the uterine cervical fluid. This is probably related to the difficulties of collecting the cervical fluid continuously and in significant amounts. In order to solve these problems, we developed a simple filter-disc-absorption method. Cervical fluid (CF) was adsorbed by a Whatman GF/B glass fiberdisc. It was then applied directly to the electrophoresis. Using this method, protein fractions and phosphorylase or lactate dehydrogenase isoenzymes in the CF of various gynecological states were investigated.
1. The CF from pregnant women in the first trimester as well as normal cycle shows weak protein fractions. As the pregnancy progress, especially after 30 gestational weeks, albumin and prealbumin levels are increased.
2. The CF obtained from patients with myoma of the uterine body and cancer of the uterine cervix contains a large amount of protein in comparison with that obtained from pregnant or normal cycle women. The electrophoretic pattern in the former cases resembles that of serum protein, especially in the cases with high protein content. However, no characteristic electrophoretic patterns which could be used to determine diseases or their severity were demonstrated.
3. Two phosphorylase bands were evident in CF from patients with the myoma and cancer. We called them the fast and the slow migrating phosphorylare bands (F-band and S-band). In the cases of cancer, the S-band was predominant, while in the cases of myoma, the F-band predominated. From these results, it was suggested that the distribution of the two phosphorylases could be used to differentiate between myoma and cervical cancer.
4. The distribution of LDH isoenzyme in the CF and in the serum of the same woman was different. The activities of LDH₁, LDH₂, LDH₃ isoenzymes were greater in the CF than in the serum. However, characteristic LDH-isoenzyme patterns of CF for typical gynecological diseases can not be determined.

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A technique of two-dimensional polyacrylamide gel electrophoresis for the separation of plasma proteins was described in detail. Human plasma proteins were separated by isoelectric focusing followed by electrophoresis in a 4 to 21% or a 4 to 30% linear gradient slab gel. Urea or SDS was not used throughout the procedure, so that the analyses of proteins in their biologically active state were possible. By this technique, human plasma protein was resolved into more than 250 spots.

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Human transferrin was purified from serum using polyacrylamide gel disc electrophoresis. The purified transferrin appeared to be free from other protein, as judged by electrophoresis on polyacrylamide gel and by immunoelectrophoresis. The purified human transferrin was resolved into a faster moving band (TF) and a slower moving band (TS) by 5.25% polyacrylamide gel electrophoresis. Analysis of the purified human transferrin by isoelectric focusing column separated this iron-transport protein into three components with isoelectric points of 5.61, 5.18 and 4.53. Electrophoresis on 5.25% polyacrylamide gel was carried out on each fraction from the column. Fractions pI 5.61 and pI 5.18 both moved with the same mobility as the slower electrophoretic component (TS). Fraction pI 4.53 moved with the same mobility as the faster electrophoretic component (TF). After recovery from the isoelectric column pI 5.18 transferrin was found to contain 0.90, 1.57 atoms of iron/mol of protein. The pI 4.53 transferrin had less than 0.05 atoms of iron/mol of protein. The pI 5.61 transferrin contained 0.51, 0.65 atoms of iron/mol of protein.

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非侵襲的なカテーテル採尿法によって得たネコ尿（0.25 μL）を試料として，ミクロ2次元電気泳動法（micro two-dimensional polyacrylamidegel electrophoresis; M2D-PAGE）による分析を行い，得られたスポットの画像分析よりニワトリ・オバルブミンを標準試料として尿中アルブミン（Alb）値を定量した．なお，スポットはウエスタンブロットおよび質量分析で同定を行い，ネコAlbであることを確認した．M2D-PAGE法による尿中Alb定量は現在獣医臨床検査で行われているヒトAlbを標準試料とした抗ヒトAlb抗体を用いた免疫比濁法に比較して高い定量値を与える測定法であることを確認した．また，慢性腎不全において国際獣医腎臓病研究グループ（The international renal interest society; IRIS）による各病期分類された第I期においてもAlbが高値を示す症例も見られることから，早期診断に有用な検査法であることが示された．加えて，正常ネコでは尿中Alb値が20 mg/dLを超える例が認められないことから，従来のカットオフ値である30 mg/dLを下回る20 mg/dLを正常値の閾値とする基準を提案する．

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PAG電気泳動のLDL分画像から変性LDLを捉え, 加齢に伴う変化や糖尿病や動脈硬化性疾患との関係について検討した. LDLの分画像の形状を, S, A, N, Dの4型に分類し, IDLやレムナントを含むN型とD型を重視した. また, LDL分画像のピークおよびLDL分画の最も速い移動部位の相対移動度から small LDLと fast β-Lpの有無を捉え, この3点を変性LDLの所見とした. LDL分画像で捉えるものとRLP-Cとは異質であり, 酸化LDLはN型・D型と fast β-Lp の異常分画に存在した. N型・D型と fast β-Lp の増加には加齢に伴う変化が観察され, 特に40歳代と50歳代との間に大きな有意差を認めた. 加齢に伴う同様の変化はT-CやLDL-Cにおいてもみられるが, その変化よりも顕著であった. 若年者 (中学2年生) のLDL分画像は, N型とD型が少ないのに対し, fast β-Lp は50歳代と同等以上の高い陽性率を示し, 酸化LDLも fast β-Lp と同様の傾向を示した. 若年者の酸化LDLは主として fast β-Lp に存在し, 成人で増加するN型・D型に存在する酸化LDLとは異質であろうと考えられた. 糖尿病のLDL分画像は, 蛋白尿 (+) 群では, N型とD型が多く, 健常者に対しても, 蛋白尿 (-) 糖尿病群に対しても有意差を認めた. 50歳代健常者と比べ, 動脈硬化性疾患群にはN型・D型と small LDLが有意に多く, fast β-Lp 陽性も増加していた.

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In order to identify urinary proteins excreted from normal subjects, we employed a new method by which molecular weight and immunochemical property of urinary proteins can be determined simultaneously. Urine was subjected to linear gradient (3∼40%) SDS-PAGE and then transferred to nitrocellulose membrane by electrophoretic blotting method. The membrane was stained with Auro Dye and blotted proteins were identified by enzyme immunoassay using specific antibodies. In this method, we identified more than 12 protein bands from urine, that is, α₂-macroglobulin, tubular epithelial antigen, Tamm-Horsfall glycoprotein, IgA, IgG, transferrin, albumin, α₁-microglobulin, L-chain (dimer, monomer), retinol-binding protein, and β₂-microglobulin. In addition, we found native forms and some fragments of immunoglobulins. In this report, we analyzed only normal subjects.
These results suggest that this method is useful for analysis not only of glomerular or tubular proteins but also tissue antigens from urine with various kinds of renal diseases.

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In my previous paper, I reported a simplified filter-disc-absorption method for analysis of cervical fluids. With this method, electrophoretic distributions of phosphorylase activities in cervical fluids were investigated.
1) Cervical fluid (CF) contained two phosphorylase isoenzymes with different mobilities: the fast moving (F-band) and the slow moving (S-band) phosphorylases. In the CF's obtained from women during a normal menstrual cycle and in normal pregnancy at the first and the middle gestational stages and from patients with myoma, the F-band isoenzyme appeared more frequently than the S-bands. In cases of which the CF contained both F-band and S-band, the former activity was much higher than the latter. In CF's of pregnant women during the last gestational stage, the S-band was more intense than the F-band. With advancement of the cancer, the S-band activity became more evident in the CF.
2) F- and S-bands of phosphorylase isoenzymes had different affinities to glycegen. The dissociation constants (K) of the isoenzymes and glycogen complexes were calculated from the results obtained by means of the affinity electrophoresis. The average K values were found to be 0.077% for the F-band and 0.029% for the S-band. These results indicated that the S-band has a higher affinity to glycogen than the F-band.

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In vitro における血清リポ蛋白の安定性について報告する. 37℃加温による血清外観の経日変化は, free では5日目, 10日目で濁りが減少し, その後, 経日的に濁りは増す. 一方, V. EやEDTAでは15日目まで変化なく, 20日目で増加した. PAGEによるリポ蛋白の経日変化については, LDL亜分画の出現, IDLの出現, LDLの陽極への移動, HDLの減少が認められた. 脂質項目の経日変化は, V. EとEDTAで安定した値が得られた. EDTAは良好なリポ蛋白変性防止効果が認められ, 血清中の金属類が酸化変性に大きく関与していることが示唆された. また同時に, in vitro において血清リポ蛋白は, 酸化変性以外にも変化することも確認された.

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36種属(ヒト,ヒト胎児,猿類21,一般動物13)のヘモグロビン(Hb)について,薄層PAG IEF法によって,人・獣血鑑別検査を試みた.その結果,Fig.1に示すような成績を得た.とくに,p-chloromercuribenzoate処理を行ったHbについてIEFを行うと,未処理Hbに比して,ヒトと動物Hbとの鑑別は容易であった(Fig.2).なお,各種属HbのpI測定値をTable 1に表示した.

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In order to detect easily the difference between human hemoglobin and animal hemoglobins, the following three techniques were examined: 1) agar gel electrophoresis, 2) chromatography on a CM-cellulose column and 3) isoelectric focusing in polyacrylamide gel.
As the results, the latter two techniques were useful for the differentiation of human hemoglobin from animal hemoglobins, and especially isoelectric focusing technique was effective with rapidity and correctness to differentiate human hemoglobin from Japanese monkey (Macaca fuscata) hemoglobin when they were treated with p-chloromercuribenzoate.

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薄層ポリアクリルアミドゲル電気泳動法を用いた血清蛋白成分の泳動分離における解像力はセルロースアセテート電気泳動法に比べはるかに優れている.しかしながら操作法の簡便さという点ではセルロースアセテート電清気泳動法に劣る.
著者らはポリアクリルアミドゲルを支持媒質とする簡易薄層電気泳動法を考案し,血清酵素蛋白成分について泳動分離条件を検討してきた.操作法の簡便化という目的のために片面がスリガラス様の粗面をもった製図用のポリエステルフィルムの上に予じめ目的に応じた緩衝液を含んだアクリルアミドゲル薄層(1mm又は2mm)を形成させて湿室に保存しておく.このようにして,薄層アクリルアミドゲルの調製操作を省くことができる.
支持シートとして,セロファン(124PDセロファン)又は片面がスリガラス様の粗面をもった製図用ポリエステルフィルムを用いた.
試料として,血清は希釈することなくゲル薄層表面に予じめ作られた溝に添加した.1mm厚さのゲル薄層には3μl,2mm厚さのゲル薄層には15μlの血清を添加する.
電極液槽用緩衝液を満した両液槽間に試料を添加したゲル薄層を置き,ゲル薄層の両端を緩衝液で湿らせた濾紙の1端を5mm程度重ねるようにのせ電極液槽との電気的連絡を行なう.
温度の上昇を防ぐために電気泳動は5℃～10℃の保冷庫内で行なった.電気泳動はゲル幅1cm当り0.5～1.0mAで,120分行なった.
泳動終了後,染色,脱色,固定したゲル薄層はガラス板上でセロファンシートの間にサンドイッチ状にはさんで室温で約2日間乾燥させ,乾燥ゲルフィルムとして保存する.
また,この簡易薄層ポリアクリルアミドゲル電気泳動法はSDSを含んだゲル薄層を用いることによって,ペプチドの分子量を簡単に,しかも正確に推定できる有利な方法である.

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