Respirometer

Abstract: Mitochondrial dysfunction is at the core of many diseases ranging from inherited metabolic diseases to common conditions that are associated with aging. Although associations between aging and mitochondrial function have been identified using mammalian models, much of the mechanistic insight has emerged from Caenorhabditis elegans. Mitochondrial respiration is recognized as an indicator of mitochondrial health. The Seahorse XF96 respirometer represents the state-of-the-art platform for assessing respiration in cells, and we adapted the technique for applications involving C. elegans. Here we provide a detailed protocol to optimize and measure respiration in C. elegans with the XF96 respirometer, including the interpretation of parameters and results. The protocol takes similar to 2 d to complete, excluding the time spent culturing C. elegans, and it includes (i) the preparation of C. elegans samples, (ii) selection and loading of compounds to be injected, (iii) preparation and execution of a run with the XF96 respirometer and (iv) postexperimental data analysis, including normalization. In addition, we compare our XF96 application with other existing techniques, including the eight-well Seahorse XFp. The main benefits of the XF96 include the limited number of worms required and the high throughput capacity due to the 96-well format.

Abstract: I Principles of Sensor Design and Operation.- I.1 Factors Influencing the Stability of Polarographic Oxygen Sensors.- I.2 Calibration and Accuracy of Polarographic Oxygen Sensors.- I.3 A Thermodynamic Consideration of Permeability Coefficients of Membranes.- I.4 Microcoaxial Needle Sensor for Polarographic Measurement of Local O2 Pressure in the Cellular Range of Living Tissue. Its Construction and Properties.- I.5 Electrolytes.- I.6 The Action of Hydrogen Sulfide on Polarographic Oxygen Sensors.- I.7 A Double-Membrane Sterilizable Oxygen Sensor.- I.8 Construction of a Polarographic Oxygen Sensor in the Laboratory.- I.9 A Polarographic Oxygen Sensor Designed for Sewage Work and Field Application.- I.10 Electronic Circuits for Polarographic Oxygen Sensors.- I.11 The Application of a Microprocessor to Dissolved Oxygen Measurement Instrumentation.- II Laboratory Applications of Polarographic Oxygen Sensors.- II.1 An Automated Multiple-Chamber Intermittent-Flow Respirometer.- II.2 The Application of Polarographic Oxygen Sensors for Continuous Assessment of Gas Exchange in Aquatic Animals.- II.3 The Twin-Flow Microrespirometer and Simultaneous Calorimetry.- II.4 Simultaneous Direct and Indirect Calorimetry.- II.5 An Automated, Intermittent Flow Respirometer for Monitoring Oxygen Consumption and Long-Term Activity of Pelagic Crustaceans.- II.6 Sealed Respirometers for Small Invertebrates.- II.7 A Method for the Simultaneous Long-Term Recording of Oxygen Evolution and Chloroplast Migration in an Individual Cell of Acetabularia.- II.8 A Respirometer for Monitoring Homogenate and Mitochondrial Respiration.- II.9 Bacterial Growth and Antibiotics in Animal Respirometry.- II.10 pO2 and Oxygen Content Measurement in Blood Samples Using Polarographic Oxygen Sensors.- II.11 Determination of the In Vivo Oxygen Flux into the Eye.- III Field Applications of Polarographic Oxygen Sensors.- III.l In Situ Measurement of Oxygen Profiles in Lakes: Microstratifications, Oscillations, and the Limits of Comparison with Chemical Methods.- III.2 In Situ Measurement of Oxygen Profiles of Sediments by Use of Oxygen Microelectrodes.- III.3 Methods for Measuring Benthic Community Respiration Rates.- III.4 In Situ Measurement of Community Metabolism in Littoral Marine Systems.- III.5 Deap-Sea Respirometry: In Situ Techniques.- Appendix A Calculation of Equilibrium Oxygen Concentration H. Forstner and E. Gnaiger.- Appendix B Calculation of pO2 in Water Equilibrated with a Mixture of Room Air and Nitrogen.- Appendix C Calculation of Energetic and Biochemical Equivalents of Respiratory Oxygen Consumption E. Gnaiger (With 1 Figure).- Appendix D The Winkler Determination H.L. Golterman (With 1 Figure).- Appendix E Symbols and Units: Toward Standardization E. Gnaiger.

Abstract: The measurement of soil carbon dioxide respiration is a means to gauge biological soil fertility. Test methods for respiration employed in the laboratory vary somewhat, and to date the equipment and labor required have somewhat limited more widespread adoption of such methodologies. The purpose of this research is to compare the results of measured soil CO2 respiration using three methods: (1) titration method; (2) infrared gas analysis (IRGA); and (3) the Solvita gel system for soil CO2 analysis. We acquired 36 soil samples from across the USA for comparison, which ranged in pH from 4.5 to 8.5, organic C from 0.8 to 4.6% and the clay content from 6 to 62%. All three methods were highly correlated with each other after 24-h of incubation (titration and Solvita r 2 = 0.82, respirometer and Solvita r 2 = 0.79 and titration versus respirometer r 2 = 0.95). The 24-h (1-day) CO2 release from all three methods was also highly correlated to both basal soil respiration (7–28 days) and cumulative 28-day CO2 respiration. An additional 24 soil samples were acquired and added to the original 36, for a total of 60 soil samples. These samples were used for calibration of the Solvita gel digital color reader results using CO2-titration results and regression analysis. Regression analysis resulted in the equation y = 20.6*(Solvita number) - 16.5 with an r 2 of 0.83. The data suggest that the Solvita gel system for soil CO2 analysis could be a simple and easily used method to quantify soil microbial activity. Applications may also exist for the gel system for in situ measurements in surface gas chambers. Once standardized soil sampling and laboratory analysis protocols are established, the Solvita method could be easily adapted to commercial soil testing labs as an index of soil microbial activity.