Cytophotometry As a Tool in Fibre Typing

Abstract

Cytophotometry is used as a technique of quantitative enzyme histochemistry based on absorbance measurements of the coloured final reaction product of an enzyme reaction in the histological section (for review, see Van Noorden and Butcher 1991). The main advantage of the cytophotometrical method is to enable measurements in defined microscopical structures such as muscle fibres (Pai et al. 1982; Reichmann and Pette 1982,1984; van der Laarse et al. 1984; Vetter et al. 1984; Blanco et al. 1988; van der Laarse et al. 1989; Blanco and Sieck 1992, this study). The levels of metabolic enzyme activities which are demonstrated histochemically by their reaction product vary in the different muscle fibres. The absorbance of the reaction product can be assumed to be proportional with its concentration in the histological section (Krug 1980). Moreover, fulfilling a set of criteria for reliability and validity of the enzyme reaction (Stoward,1980, van Noorden and Frederiks 1992), the absorbance of the reaction product is additionally proportional with enzyme activity and can serve as a measure for that. Direct comparison of cytophotometry and biochemistry is possible by calibration of cytophotometrical values (van Noorden and Frederiks 1992). In this way, data of both methods can be expressed as moles of substrate converted per unit time per unit wet weight of tissue. However, for many investigations such as muscle fibre analysis, absolute values of enzyme activities are not necessary. It is rather the comparison of enzyme activities in different structures or under changing experimental conditions that is of interest. In these cases, one can use enzyme activities as percentages related to a reference value, and calibration procedures are unnecessary.