生物物理化学
Online ISSN : 1349-9785
Print ISSN : 0031-9082
ISSN-L : 0031-9082
19 巻, 2 号
選択された号の論文の10件中1~10を表示しています
  • 太中 弘
    1974 年 19 巻 2 号 p. 97-100,1
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
  • David Gitlin
    1974 年 19 巻 2 号 p. 101-112
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
  • 内木 詢一
    1974 年 19 巻 2 号 p. 113-121
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
    The rabbit liver, heart, and muscle extracts showed isoelectric focusing peak patterns of LDH (Ampholine pH 3.5-10), which considerably differed from the LDH zymograms characteristic to those tissues. Ampholine did not give any significant influence on enzymic activity and electrophoretic mobility (agar gel plate) of the five fractions of purified LDH isozymes. Isoelectric focusing of LDH band 1 (H4) produced an acidic peak but with a very poor recovery of activity (1-2%), while that of LDH band 5 (M4) gave an alkaline peak with a good recovery of activity (88%). LDH bands 2 (H3M), 3 (H2M2), and 4 (HM3) exhibited multiple isoelectric focusing peaks, of which an alkaline peak was dominant. Isoelectric focusing of the DANSylated LDH isozyme, band 3, revealed 3 to 4 fluorescent protein peaks not corresponding to the activity peaks. Dissociaton and partial reactivation of LDH (tetramer), accompanying aggregation-denaturation (inactivation), especially in acidic pH, are responsible for producing the modified activity peak pattern of LDH isozymes in the isoelectric focusing experiments.
  • 高原 喜八郎
    1974 年 19 巻 2 号 p. 123-128
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
    A new method was developed for the whole blood direct determination of blood sugar by an ultrahighly sensitive ortho-toluidine reagent (OT-37).
    The sensitivity of OT-37 was 2.6 fold higher than the regular reagents. The procedure for the determination was as follows. 10μl of whole blood was mixed with 5ml of OT-37 by blowing out from a capillary pipette. The blood hemolyzed immediately on mixing. Absorbance at 565nm (Eb) was read and the tube was heated at 100°C for 3 minutes and 30 seconds. After cooling the tube in running water, absorbance at 565nm (Ea) was read again. The difference between the two readings, ΔE=Ea-Eb, was found to be proportional to the glucose concentration.
    In contrast with the negative interference caused by hemoglobin iron at the regular wave length of determination (635nm), no interference was observed at 565nm. Addition of 10μl of 10mg/ml glutathione, either reduced or oxidized, gave no effect on readings. Positive interference caused by NAD or NADH was found to be cancelled by coexisting hemoglobin.
    Recovery tests and comparison with the values obtained by conventional methods gave satisfactory results.
    The new method seemed, therefore, to be satisfactory and useful especially in emergency determinations.
  • 宮城 芳得
    1974 年 19 巻 2 号 p. 129-137
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
    The staining ability of 5 dyes was compared by staining a known amount of human serum albumin spotted on a sheet of Separax, eluting the protein-bound dye, and measuring absorbancy at respective peak wave lengths. The relative color intensity of G-250 was much higher than that of R-250, Ponceau 3R, Amidoblack 10B or Procion blue. The colorimetric value of G-250 binding to serum protein fractions considerably fluctuated because of an unavoidable inaccuracy of spotting of the protein solution onto Separax. However, the relative proportion of protein fractions (expressed in percentage) showed a reasonable variation, especially when serum was diluted 5-fold with saline. The similar tendency was obtained when the stained protein fractions were measured using densitometer. A few examples of the G-250 staining method applied to paper electrophoresis of cerebrospinal fluid and urinary proteins were presented.
  • 大槻 真, 佐伯 進, 尤 芳才, 近藤 勤, 馬場 茂明
    1974 年 19 巻 2 号 p. 139-142
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
    Horizontal thin-layer polyacrylamide gel electrophoresis proved to provide a good resolution of urine amylase isozymes, with up to eight bands. In order to determine the source of amylase isozymes in urine, comparative studies were made on the electrophoretic mobilities of amylase bands in urine, salivary gland and pancreas homogenates obtained under the same condition. The results suggested that the amylase isozymes contained in urine consisted of those originated in pancreas and salivery glands. Storage of urine at 4°C and -18°C for 6 months resulted in no change of the mobilities of urine amylase isozymes. From these results, it seemed to be reasonable to determine the origin of each amylase isozyme on the basis of the electrophoretic mobility. Heat treatment of urine and saliva at 56°C for 6 hours caused no or slight reduction in amylase activity. The amylase activity of pancreatic juice, however, was completely inactivated by heat treatment at 56°C for 60 minutes. The mobility of each amylase isozyme did not change by the heat treatment, while, the activity of the isozyme AmyU-1 (pancreatic origin) showed marked reduction. This behavior suggests a possible physicochemical difference between the pancreatic and salivary amylases.
  • 有好 邦夫
    1974 年 19 巻 2 号 p. 143-150
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
    A vertical polyacrylamide gel slab electrophoresis apparatus, which employed a discontinuous buffer system, and spacer gel 5mm thick at the upper portion, 1mm at the lower portion and running gel 1mm in thickness, was devised.
    The increased thickness of the upper portion of the spacer gel and the discontinuous buffer system made it possible to apply a large quantity of dilute sample and to obtain a sharp electrophoretogram similar to that of disc electrophoresis.
    The thinness of the running gel made it possible to use ultraviolet method for detecting isozyme.
    Using this method, human lactate dehydrogenase isozymes and pyruvate kinase isozymes were demonstrated in sharp bands. Furthermore, purification of human erythrocyte pyruvate kinase was performed and purity of the enzyme in each step of purification was examined with simultaneous comparison of protein and enzyme stain. These results confirmed that this apparatus was useful for the study of dilute enzyme preparations especially of unstable enzyme such as pyruvate kinase.
  • 古賀 俊逸, 平山 千里, 井林 博
    1974 年 19 巻 2 号 p. 151-155
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
    Quantitative analysis of serum high density lipoprotein (HDL) was carried out by the rocket immunoelectrophoresis using monovalent antiserum against HDL. The antisera were prepared in rabbits injecting each of two major subunit components of apoHDL. Subunit peptides of apoHDL were fractionated from normal human apoHDL by a gel-filtration through Sephadex G-200 suspended in 8M urea (Scanu et al., Biochemistry 8, 3306 (1967)). Two major subunit peptides, apoHDL (III) and apoHDL (IV) have been known immunochemically distinct. These two subunit peptides are equivalent to A-I and A-II peptide or apoLp-Gln-I and apoLp-Gln-II. Single line of precipitation was observed by rocket immunoelectrophoresis of serum samples on an agarose-gel plate containing eaah of anti-apoHDL (III) or anti-apoHDL (IV) serum. The linearity of the standard curve and the reproducibility of the determinations were good.
    HDL concentration in normal human subjects at age 18-22 years old revealed 106±18 and 121±8 units for male and female, respectively (100 units for reference serum). Concentration of HDL in various kinds of liver diseases were 40±30 units for acute hepatitis (7 patients), 92±20 units for chronic hepatitis inactive form (18 patients), 85±21 units for chronic hepatitis active form (8 patients), 80±31 units for cirrhosis (25 patients) and 50±33 units for hepatoma (8 patients). All of them decreased significantly from normal controls (109±19 units for 24 subjects). No significant differences were observed in the results of HDL determination whether measured by anti-apoHDL (III) or anti-apoHDL (IV) serum.
  • 125I標識ラット血清アルブミンおよびγ-グロブリンのラット肝臓,脾臓組織へのin vitroでのとりこみ
    小松 信三, 柴田 泰生
    1974 年 19 巻 2 号 p. 157-163
    発行日: 1975/02/25
    公開日: 2009/03/31
    ジャーナル フリー
    健常動物での血清alb,γ-glob濃度の恒常性をそれら蛋白産生のfeedback調節によって解釈しようという試みは,in vivo実験で証明できず,高濃度時の異化促進などによって説明されている.しかし,最近家兎網状赤血球でのヘモグロビン産生,マウス骨髄腫細胞での免疫glob産生について,cell-free培養で完成蛋白(前者ではグロビンα鎖,後者では免疫グロブリン)が,合成過程におけるアミノ酸配列順序のよみとりの段階を抑制または調節することが報告されている.このようなin vivoin vitroの結果の相違を考えるとき,まず組織による血清蛋白のとりこみ如何が問題となる.in vivoではラットで肝臓などによる131I標識albの異化が報告され,pinocytosisに始まる細胞内異化順路の推論も出されている.
    われわれはラット血清albならびにγ-globを125I標識して,ラット肝,脾組織試料とin vitro孵置し,これら蛋白の組織との相互作用をしらべた.組織ミンス200mg当たり5mg程度の標識蛋白をin vitro投与すると37℃,1時間培養後,1回洗浄した組織部分にかなりの放射活性が移行する(albの場合,投与L量の6.8%(肝),8.0%(脾),γ-globの場合.11.5%(肝),5.6%(脾)).この移行は孵置時間とともに増大し,細胞分画を行なって検討したところ,放射活性の細胞内分布は臓器種,蛋白種別に異なり,また,時間別に変化した.さらに孵置後の細胞の上清分画の電気泳動分析で,蛋白性125I放射活性の大部分が投与標識蛋白と等しい易動度を示した.以上からin vitroでの放射活性の組織への移行は吸着によるものでなく,組織による標識蛋白のとりこみと推定し得る.
    同時にその際,両組織の蛋白種別とりこみの程度に差が見られた.Albとりこみ量対γ-globとりこみ量の比を仮にA対G比として表現すると,肝臓組織ではこの値が1時間孵置で0.6,脾臓組織では1.6となり,両組織ともそれぞれの産生蛋白種を他より少なくとりこむ結果が得られた.一方,両組織よりミクロゾームを調製して,cellfree条件で標識蛋白との孵置を行なうと,30分孵置でA対G比は肝臓の場合1.04,脾臓の場合0.75と,それぞれ自己の産生蛋白種を多く結合する傾向が示された.これはマウス骨髄腫蛋白が骨髄腫細胞のmRNAと特異的に結合するとの他家の報告と並行する結果であり,細胞下レベルでは自己産生蛋白種との親和性が強い.
    以上から,肝臓および脾臓の組織に蛋白種弁別,とりこみ抑制機構の存在が考えられ,これがまた他家によるin vivo実験でのfeedback調節の証明不成功につながったと推定する.
  • 1974 年 19 巻 2 号 p. e1
    発行日: 1974年
    公開日: 2009/03/31
    ジャーナル フリー
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