Further experimental studies on the horseradish peroxidase-oxidase reaction

1992 ◽  
Vol 96 (18) ◽  
pp. 7338-7342 ◽  
Author(s):  
Marjorie S. Samples ◽  
Yu Fen Hung ◽  
John Ross
Keyword(s):  
Horseradish Peroxidase ◽  
Experimental Studies ◽  
Oxidase Reaction

scholarly journals Kinetic changes in activity of HR-peroxidase, induced by very low doses of phenol

2021 ◽  
Vol 7 (23) ◽  
pp. 48-55
Author(s):  
Elzbieta Malarczyk
Keyword(s):  
Horseradish Peroxidase ◽  
Maximum Activity ◽  
Low Doses ◽  
Sensitive Detector ◽  
High Dilution ◽  
Palabras Clave ◽  
High Dilutions ◽  
Sinusoidal Pattern ◽  
Activity Dependent ◽  
Oxidase Reaction

The allosteric protein of horseradish peroxidase (HRP) shows two main types of activity, peroxidase and oxidase, depending on the kind of low molecular effectors. The effects of very low doses of phenol, prepared by successive dilutions in water or in 75% ethanol, on initial HRP activity in oxidation of o-dianisidine or luminol were tested in a systematic manner by colorimetric and luminometric methods. Results showed that phenol dilutions, including those below Avogadro’s number, could activate or inhibit HRP in peroxidase and oxidase type reactions with a sinusoidal pattern. Km values for the studied substrates changed parallel to HRP peroxidase/oxidase activity and the maximum activity in the peroxidase reaction corresponded to the minimum activity in the oxidase reaction and vice versa. The effect also depended on the type of dilutor. The observations of the peroxidase/oxidase oscillations in the sinusoidal pattern of HRP activity, dependent on the rate of phenol dissolution and the time of preincubation, point out to the conclusion that HRP might be a good model for high dilutions research. The experiments provide strong evidence that horseradish peroxidase (HRP) is a very sensitive detector of subtle changes in the concentration of phenol used as a cofactor in the peroxidase/oxidase reaction. Keywords: HR-peroxidase, peroxidase-oxidase, phenol, hormesis, homeopathy, high dilutions.   Mudanças cinéticas na atividade da HR-peroxidade induzidas por doses muito baixas de fenol Resumo A proteína alostérica da peroxidase do rabano (HRP) mostra dois tipos principais de atividade, peroxidase e oxidase, de acordo com o tipo de efetores de baixa molecularidade. Os efeitos de doses muito baixas de fenol, preparadas através de diluições sucessivas em água ou etanol 75% na atividade inicial da HRP sobre a oxidação da o-dianisidina ou luminol for testados de modo sistemático através de métodos colorimétricos e luminométricos. Os resultados mostram que as diluições de fenol, incluindo aquelas por baixo do número de Avogadro, foram capazes de ativar ou inibir a HRP em reações de tipo peroxidade e oxidase com um padrão sinusoidal. os valores Km dos substratos estudados variaram paralelamente à atividade peroxidase/oxidase da HRP; a atividade máxima da reação peroxidase correspondeu à atividade mínima na reação oxidase e vice-versa. O efeito também se mostrou dependente do tipo do solvente. A observação das oscilações sinusoidais na atividade da HRP, dependentes da taxa de dissolução do fenol e do tempo de pré-incubação, permitem concluir que a HRP pode ser um bom modelo na pesquisa das altas diluições. Os experimentos oferecem fortes evidéncias a favor da HRP como detector muito sensível de mudanças mínimas na concentração do fenol, utilizado como cofator na reação peroxidase/oxigenase. Palavras-chave: HR-peroxidase, peroxidase-oxidase, fenol, hormese, homeopatia, altas diluições.   Cambios cinéticos en la actividad de la HR-peroxidasa inducidos por dosis muy bajas de fenol Resumen La proteína alostérica de la peroxidasa del rábano (HRP) muestra dos tipos principales de actividad, peroxidasa y oxidasa, dependiendo del tipo de efectores de baja molecularidade. Los efectos de doses muy bajas de fenol, preparadas mediante diluciones sucesivas en agua o etanol al 75% sobre la actividad inicial de la HRP sobre la oxidación de o-dianisidina o luminol fueron testados de modo sistemático mediante métodos colorimétricos y luminométricos. Los resultados muestran que las diluciones de fenol, incluyendo aquellas abajo del número de Avogadro, pudieron activar o inhibir la HRP en reacciones de tipo peroxidasa y oxidasa con un patrón sinusoidal. Los valores Km de los sustratos estudiados variaron paralelamente a la actividad peroxidasa/oxidasa de la HRP; la actividad máxima de la reacción peroxidasa correspondió a la actividad mínima en la reacción oxidasa y viceversa. El efecto también se mostró dependiente del tipo de solvente. La observación de las oscilaciones sinusoidales en la actividad de la HRP, dependientes de la tasa de disolución del fenol y del tiempo de preincubación, llevan a concluir que la HRP puede ser un buen modelo para la investigación de las altas diluciones. Los experimentos ofrecen fuertes evidencias a favor de la HRP como detector muy sensible de cambios mínimos en la concentración de fenol, utilizado como cofactor en la reacción peroxidasa/oxigenasa. Palabras-clave: HR-peroxidasa, oxidasa-peroxidasa, fenol, hormesis, homeopatía, altas diluciones.   Correspondence author: Elzbieta Malarczyk, [email protected] How to cite this article: Malarczyk E. Kinetic changes in the activity of HR-peroxidase induced by very low doses of phenol. Int J High Dilution Res [online]. 2008 [cited YYYY Mmm DD]; 7(23): 48-55. Available from: http://journal.giri-society.org/index.php/ijhdr/article/view/37/349.  

Effects of Vincristine on Endothelial Microtubules in the Spinal Cord

1976 ◽  
Vol 34 ◽  
pp. 58-59
Author(s):  
John L. Beggs ◽  
John D. Waggener ◽  
Wanda Miller
Keyword(s):  
Spinal Cord ◽  
Biological Activity ◽  
Horseradish Peroxidase ◽  
Transport Mechanism ◽  
Vasogenic Edema ◽  
Vesicular Transport ◽  
Close Proximity

Microtubules (MT) are versatile organelles participating in a wide variety of biological activity. MT involvement in the movement and transport of cytoplasmic components has been well documented. In the course of our study on trauma-induced vasogenic edema in the spinal cord we have concluded that endothelial vesicles contribute to the edema process. Using horseradish peroxidase as a vascular tracer, labeled endothelial vesicles were present in all situations expected if a vesicular transport mechanism was in operation. Frequently,labeled vesicles coalesced to form channels that appeared to traverse the endothelium. The presence of MT in close proximity to labeled vesicles sugg ested that MT may play a role in vesicular activity.

Comparative strengths and weaknesses of peroxidase and colloidal gold in pre- and post-embedding immunolabeling techniques

1987 ◽  
Vol 45 ◽  
pp. 1002-1005
Author(s):  
D. R. Abrahamson ◽  
P. L. St.John ◽  
E. W. Perry
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Light Microscopy ◽  
Colloidal Gold ◽  
Light Microscope ◽  
Ultrastructural Localization ◽  
The Other ◽  
Quantitative Studies ◽  
Dense Suspension ◽  
Antigen Localization

Antibodies coupled to tracers for electron microscopy have been instrumental in the ultrastructural localization of antigens within cells and tissues. Among the most popular tracers are horseradish peroxidase (HRP), an enzyme that yields an osmiophilic reaction product, and colloidal gold, an electron dense suspension of particles. Some advantages of IgG-HRP conjugates are that they are readily synthesized, relatively small, and the immunolabeling obtained in a given experiment can be evaluated in the light microscope. In contrast, colloidal gold conjugates are available in different size ranges and multiple labeling as well as quantitative studies can therefore be undertaken through particle counting. On the other hand, gold conjugates are generally larger than those of HRP but usually can not be visualized with light microscopy. Concern has been raised, however, that HRP reaction product, which is exquisitely sensitive when generated properly, may in some cases distribute to sites distant from the original binding of the conjugate and therefore result in spurious antigen localization.

3-D reconstruction and analysis of mammalian mitotic spindle structure

1992 ◽  
Vol 50 (1) ◽  
pp. 578-579
Author(s):  
Kent McDonald ◽  
David Mastronarde ◽  
Rubai Ding ◽  
Eileen O'Toole ◽  
J. Richard McIntosh
Keyword(s):  
Cross Sections ◽  
Mitotic Spindle ◽  
Experimental Studies ◽  
Video Camera ◽  
Three Dimensions ◽  
Mitotic Spindles ◽  
Black And White ◽  
Reconstruction Methods ◽  
Spindle Structure ◽  
Tissue Culture Line

Mammalian spindles are generally large and may contain over a thousand microtubules (MTs). For this reason they are difficult to reconstruct in three dimensions and many researchers have chosen to study the smaller and simpler spindles of lower eukaryotes. Nevertheless, the mammalian spindle is used for many experimental studies and it would be useful to know its detailed structure.We have been using serial cross sections and computer reconstruction methods to analyze MT distributions in mitotic spindles of PtK cells, a mammalian tissue culture line. Images from EM negatives are digtized on a light box by a Dage MTI video camera containing a black and white Saticon tube. The signal is digitized by a Parallax 1280 graphics device in a MicroVax III computer. Microtubules are digitized at a magnification such that each is 10-12 pixels in diameter.

Unlabeled Antibody Immunocytochemistry Applied to Murine Mammary Tumor Cells

1975 ◽  
Vol 33 ◽  
pp. 390-391
Author(s):  
Wm. J. Arnold ◽  
J. Russo ◽  
H. D. Soule ◽  
M. A. Rich
Keyword(s):  
Horseradish Peroxidase ◽  
Mammary Tumor ◽  
Covalent Binding ◽  
Petri Dish ◽  
Gamma Chain ◽  
Mammary Tumor Virus ◽  
Mammary Tumor Cells ◽  
Murine Mammary Tumor ◽  
Tumor Virus ◽  
Unlabeled Antibody

Our studies of mammary tumor virus have included the application of the unlabeled antibody enzyme method of Sternberger to mammary tumor derived mouse cells in culture and observation with an electron microscope. The method avoids the extravagance of covalent binding of indicator molecules (horseradish peroxidase) with precious antibody locator molecules by relying instead upon specific antibody-antigen linkages. Our reagents included: Primary Antibody, rabbit anti-murine mammary tumor virus (MuMTV) which was antiserum 113 AV-2; Secondary Antibody, goat anti-rabbit IgG gamma chain (Cappel Laboratories); andthe Indicator, rabbit anti-horseradish peroxidase - horseradish peroxidase complex (PAP) (Cappel Labs.). Dilutions and washes were made in 0.05 M Tris 0.15 M saline buffered to pH 7.4. Cell monolayers, after light fixation in glutaraldehyde, were incubated in place by a protocol adapted from Sternberger and Graham and Karnovsky, then embedded by our usual method for monolayers. Reagents were confined to specific areas by neoprene 0-rings (Parker Seal Co.) reducing the amount of reagent needed to 50 microliters, 1/6th of that required to wet a 35 mm petri dish.

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