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  •       Subsampling

In Constant Volume Respirometry (CVR), or 'closed system' respirometry, the organism is placed in a sealed chamber, and over time its respiration changes the gas concentrations in the chamber.   You measure rates of gas exchange by determining gas concentrations at the start and end of a period of measurement, and then using the cumulative difference in concentrations and the elapsed time to compute the average rate of change*.

For small organisms (e.g., insects; bird or reptile eggs), it is convenient to use large-volume syringes with shutoff valves as chambers -- these can be used to inject gas samples directly into the analysis system, as shown below (be sure to avoid crushing your organism during the injection process!!).   For big animals, use a big (e.g., 60 cc) syringe to suck a sample out of the (much larger) sealed animal chamber at the end of the measurement period.

For CVR, I have settled on the following tubing arrangement for quick and accurate measurements of gas concentrations, particularly for experiments involving numerous simultaneous samples (e.g., you put a large number of test subjects, each in its own chamber, into an incubator at about the same time, and then measure them all in rapid sequence after the experimental interval has elapsed):

Note that for simplicity, this diagram does not show the gas scrubbing agents -- desiccants; CO₂ absorbants -- that are normally part of a gas analysis system.   These would fit between the injection site (T-junction) and the analyser(s).

Be sure the flowrate through the analyzer is low:   perhaps 20-30 ml/min -- and even lower if your injection volume is less than 50-60 ml.   Inject your samples fairly rapidly.   As long as the opposite end of the tubing is open to the atmosphere, the injected volume will 'backfill' the tube and then be pulled evenly through the analyzer.   This should yield a smooth peak with a stable flat 'top' indicating the end-period gas concentration.   Note that there will likely be a sharp transient peak in the O₂ or CO₂ record resulting from the pressure spike during the injection.   Ignore this during analysis.

*This page shows the standard CVR approach, but in some cases it is possible to continuously monitor gas concentration within the closed chamber as the animal's respiration affects it over time (e.g., with an oxygen electrode or a recirculation circuit incorporating the analyer).   This would be a decline in O2 concentration, or an increase in CO2 concentration over time.  In either case, if the data are clean enough, continuous (time-specific) rates of gas exchange may be computed by taking the derivative of gas concentrations over time.  In that scenario, the plumbing arrangement shown on this page is irrelevant.