Related Experiment Video
Updated: May 23, 2026
High-resolution Respirometry to Assess Mitochondrial Function in Permeabilized and Intact Cells
Published on: February 8, 2017
Effects of digitonin on mitochondrial and lysosomal fractions from renal cortex
Abstract:
Digitonin was studied as a means of purifying mitochondrial fractions which were contaminated by lysosmes and other membranous structures. At concentrations which did not cause loss of mitochondrial enzymes, digitonin released into the supernatant significant amounts of marker enzymes for lysosomes, plasma membranes, and other cytoplasmic organelles. Acid glycosidases followed differing patterns of release that may reflect the heterogenous organization of the lysosomal enzyme-membrane complex. While there are limitations to the digitonin method, studies of certain aspects of mitochondrial function may benefit from the digitonin pretreatment step.
Insights
Digitonin effectively purifies mitochondrial fractions by releasing contaminants like lysosomes without damaging essential mitochondrial enzymes. This method aids specific mitochondrial function studies despite some limitations.
Area of Science:
- Biochemistry
- Cell Biology
- Mitochondrial Research
Background:
- Mitochondrial purification is crucial for studying mitochondrial function.
- Contamination by other organelles, such as lysosomes, complicates mitochondrial isolation.
- Digitonin is a saponin used for membrane permeabilization.
Purpose of the Study:
- To evaluate digitonin as a tool for purifying mitochondrial fractions.
- To assess the impact of digitonin on mitochondrial enzymes and contaminants.
Main Methods:
- Digitonin treatment of cellular fractions.
- Assay of marker enzymes for mitochondria, lysosomes, plasma membranes, and other organelles in supernatant and pellet fractions.
- Analysis of acid glycosidase release patterns.
Main Results:
- Digitonin released significant amounts of lysosomal, plasma membrane, and other organelle marker enzymes at non-damaging concentrations for mitochondrial enzymes.
- Acid glycosidases exhibited varied release patterns, suggesting complex lysosomal organization.
- The digitonin method demonstrated potential for reducing contamination in mitochondrial preparations.
Conclusions:
- Digitonin can selectively permeabilize membranes of contaminating organelles without compromising mitochondrial integrity.
- The differential release of enzymes highlights the utility of digitonin in specific mitochondrial research.
- While not a universal solution, digitonin pretreatment offers benefits for certain mitochondrial studies.
