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Assessment of lysosomal function by quantitative histochemical and cytochemical methods

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Quantitative histochemistry and cytochemistry enables a direct link to be made between metabolic functions such as the activity of lysosomal enzymes and the morphology of a tissue or a type of cell. Several approaches exist such as microchemistry based on (bio)chemical analysis of a single cell or a small piece of tissue dissected from a freeze-dried section. This technique has been routinely used for prenatal diagnosis of inherited enzyme defects and especially of lysosomal storage diseases. Other approaches are cytofluorometry or cytophotometry, which are based on the principle that a fluorescent or coloured final reaction product is precipitated at the site of the enzyme. The amount of final reaction product is analysed per cell or per unit volume of tissue using either a microscope cytofluorometer or flow cytometer for fluorescence measurements or an image analysing system or scanning and integrating cytophotometer for absorbance measurements.

In principle, fluorescence methods are to be preferred over chromogenic methods because they are more sensitive and enable multiparameter analysis. However, only a limited number of fluorogenic methods are at hand that give a final reaction product which is sufficiently water-insoluble to guarantee good localisation. The best results have been obtained with methods based on naphthol AS-TR derivatives and with methods for the demonstration of protease activity using methoxynaphthylamine derivatives as substrates and 5′-nitrosalicylaldehyde as coupling reagent. Chromogenic methods are far better with respect to localisation properties and, therefore, most commonly used for quantitative histochemical analysis of lysosomal enzyme activities. Besides the measurement of enzyme reactions in tissues and cells, chromogenic methods have been applied for the analysis of kinetic parameters of lysosomal enzymesin situ which could be a better reflection of enzyme kineticsin vivo than those obtainedin vitro with biochemical means in diluted solutions. Chromogenic methods have also been used in the lysosomal fragility test which is based on the lag phase occurring when a lysosomal enzyme reaction is analysed against time. The duration of the lag phase is a parameter for the stability of the lysosomal membrane and is affected by toxic compounds or under pathological conditions. This paper reviews briefly fundamental aspects and applications of quantitative histochemical and cytochemical methods in the study of lysosomes.

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  1. Laboratory of Cell Biology and Histology, University of Amsterdam, Academic Medical Center, Meibergdreef 15, 1105 AZ, Amsterdam, The Netherlands

    Cornelis J. F. Van Noorden

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Van Noorden, C.J.F. Assessment of lysosomal function by quantitative histochemical and cytochemical methods. Histochem J 23, 429–435 (1991). https://doi.org/10.1007/BF01041372

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