The principle of determining relative enzyme activities by comparative kinetic microphotometryin situ

Pette, Dirk; Reichmann, Heinz

doi:10.1007/bf01753353

Cited by 8 publications

(3 citation statements)

References 15 publications

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“…Thus far, image analysis was used as an automated version of a cytophotometer to perform endpoint measurements after a set incubation time (20)(21)(22)(23)(24)(25)(26). However, specific enzyme reactions may or may not be linear in time (11,12,27,28). Linearity may be disturbed when the product that is formed begins to inhibit the enzyme reaction.…”

Section: Introductionmentioning

confidence: 99%

Image analysis and image processing as tools to measure initial rates of enzyme reactions in sections: distribution patterns of glutamate dehydrogenase activity in rat liver lobules.

Jonker

Geerts²,

Charles³

et al. 1995

J Histochem Cytochem.

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To analyze regional differences in the activity of glutamate dehydrogenase in rat liver in situ, we developed an image recording and processing system for monitoring the formation of a colored final reaction product in time. All absorbance measurements of test and control reactions in time in consecutive sections were used to fit the data to a quadratic curve, with the derivative at t = 0 representing the initial velocity of formazan formation. The images of sections incubated for test and control reactions were topographically matched with an affine transformation using the positions of vessels as fiducials. Specific enzyme activity was calculated by subtracting the coefficients representing the initial

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Section: Introductionmentioning

confidence: 99%

Image analysis and image processing as tools to measure initial rates of enzyme reactions in sections: distribution patterns of glutamate dehydrogenase activity in rat liver lobules.

Jonker

Geerts²,

Charles³

et al. 1995

J Histochem Cytochem.

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“…Contrary to commonly used single point ("end point") measurements (Martin et al 1986;Blanco et al 1988;Powers et al 1993;Nakae and Stoward 1995;Sieck et al 1995), kinetic microphotometric determination of local enzyme activities in tissue sections is based on the evaluation of maximum reaction rates (Nolte and Pette 1972;Wimmer 1979, 1980;Lomax et al 1989;Old and Johnson 1989;Pette and Reichmann 1989;Nakae and Stoward 1995). As has been outlined elsewhere (Leeuw and Pette 1994), measurements of reaction rates by integrating microdensitometry within the same section largely exclude artefacts due to distributional error and non-uniform illumination (Goldstein 1981).…”

Section: Methodological Aspectsmentioning

confidence: 99%

Time-dependent increase of succinate dehydrogenase activity in low-frequency stimulated rabbit muscle: a comparison between microphotometric and biochemical methods

Škorjanc

Heine²,

Pette³

1997

Histochemistry and Cell Biology

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We present an improved method for microphotometric measurement of enzyme activity in muscle fibres by determining maximum reaction rates using computer-assisted image analysis. The method was used to determine absolute and relative activities of succinate dehydrogenase (SDH) in 4801 whole-fibre cross-sections of rabbit tibialis anterior muscles stimulated at low frequency (10 Hz) for different time periods of up to 50 days. Measurements were performed on cross-sections of composite blocks from stimulated and contralateral control muscles. The validity of the method was checked by determining SDH activity in homogenates of the same muscles using a standard photometric assay. Both methods yielded similar results for the time-dependent increases of SDH activity in response to chronic low-frequency stimulation. Significant increases in catalytic activity were detected by the two methods only in muscles stimulated for longer than 8 days. According to homogenate measurements, overall SDH activity was 7.4-fold elevated in 50-day-stimulated total muscles. Depending on whether or not measurements were corrected for the so-called nothing-dehydrogenase activity, the average increase in microphotometrically determined SDH activity amounted to approximately 8-fold or 10-fold, respectively. Microphotometry revealed pronounced scattering of SDH activities within the fibre populations of both normal and stimulated muscles. The heterogeneity of the fibre population with regard to SDH activity increased in long-term stimulated muscles ranging between 5-fold and 15-fold elevations.

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“…For SDH activity, the procedure of Pette and Reichmann (1989) was selected. The activity is based on the OD obtained as an endpoint terminated after the reaction was allowed to run for 10 min.…”

Section: Tissue Sampling and Preparationmentioning

confidence: 99%

A pilot study to determine whether differences exist in histochemical properties between the trapezius and extensor carpi radialis brevis muscles in women with work-related myalgia

Green

Ranney

Burnett

et al. 2014

Can. J. Physiol. Pharmacol.

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To investigate fibre-type abnormalities in women with work-related myalgia (WRM), tissue samples were extracted from their trapezius (TRAP) and the extensor carpi radialis brevis (ECRB) muscles and compared with healthy controls (CON). For the ECRB samples (CON, n = 6; WRM, n = 11), no differences (P > 0.05) were found between groups for any of the properties examined, namely fibre-type (I, IIA, IIX, IIAX) distribution, cross-sectional fibre area, capillary counts (CC), capillary to fibre area ratio, and succinic dehydrogenase activity. For the TRAP samples (CON, n = 6; WRM, n = 8), the only difference (P < 0.05) observed between groups was for CC (CON > WRM), which was not statistically significant (P > 0.05) when age was used a covariant. A comparison of the properties of these 2 muscles in the CON group indicated a higher (P < 0.05) and lower (P < 0.05) percentage of type I and type IIA fibres, respectively, in the TRAP as well as higher (P < 0.05) CC, which was not specific to fibre type. These preliminary results suggest that the properties employed to characterize fibre types do not differentiate CON from WRM for either the TRAP or ECRB. As a consequence, the role of inherent fibre-type differences between these muscles in the pathogenesis of WRM remains uncertain.

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The principle of determining relative enzyme activities by comparative kinetic microphotometryin situ

Abstract: SummaryThe principles by which the relative activities of enzymes in situ can be determined by microphotometry are reviewed.

Cited by 8 publications

References 15 publications

Image analysis and image processing as tools to measure initial rates of enzyme reactions in sections: distribution patterns of glutamate dehydrogenase activity in rat liver lobules.

Image analysis and image processing as tools to measure initial rates of enzyme reactions in sections: distribution patterns of glutamate dehydrogenase activity in rat liver lobules.

Time-dependent increase of succinate dehydrogenase activity in low-frequency stimulated rabbit muscle: a comparison between microphotometric and biochemical methods

A pilot study to determine whether differences exist in histochemical properties between the trapezius and extensor carpi radialis brevis muscles in women with work-related myalgia

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