**2. Relevant biological processes**

The term "*cellular bioenergetics*" describes the biochemical processes involved in energy metabolism (energy conversion or transformation), while the term "*cellular respiration*" describes delivery of O2 to the mitochondria, the breakdown of reduced metabolic fuels with passage of electrons to O2, and the resulting synthesis of ATP. Impaired respiration thus implies any abnormality involving cellular bioenergetics, including glycolysis. The term "*apoptosis*" describes cellular mechanisms responsible for initiating and executing cell death. The initiation step requires a leakage of cytochrome c from the mitochondrial intermembrane to the cytosol. In the cytosol, cytochrome c binds to the apoptotic protease activating factor-1 (Apaf-1), triggering the caspase cascade (a series of cysteine, aspartatespecific proteases). Caspase activation executes mitochondrial dysfunction (Nicholson et al., 1997). This mitochondrial perturbation involves opening the permeability transition pores (accelerating oxidations in the mitochondrial respiratory chain) and collapsing the

Phosphorescence Oxygen Analyzer as a Measuring Tool for Cellular Bioenergetics 239

(http://www.mccdaq.com/daq-software/universal-library.aspx). It allows direct reading from the PCI-DAS 4020/12 I/O Board (http://www.mccdaq.com/pci-dataacquisition/PCI-DAS4020-12.aspx). The software utilizes a relational database that stores experiments, pulses and pulse metadata, including slopes. Pulse identification is performed by detecting 10 phosphorescence intensities above 1.0 volt (by default). Peak identification is performed by the program which detects the highest 10% data points of a pulse and chooses the point in the group that is closest to the pulse's decay curve. Depending on the sample rate, a minimum number of data points per pulse is set and used as a cutoff to remove

Main advantages of the developed program over commercially available packages (e.g., DASYLab™ or TracerDAQ™) are provision of full control and customization of the data acquisition, storage and analysis. The choices of VB6 and Access as programming and storage environments are due to their availability, simplicity, widespread use and VB6 components that read directly from the PCI card made available by Measurement Computing. Table 1 displays identified tasks of the program. Fig. 1 shows a picture of the data acquisition system and the developed software program. Fig. 2 shows a reaction vial.

invalid pulses with too few data points (Shaban, 2010).

Experiment identification (title, date, time and sample rate)

Reading directly from the PCI card at the fastest possible rate

Calculating the exponential decay rate (1/) and lifetime () of each pulse

Storing each pulse data points, along with the peak, decay and lifetime values

Ability to remove erroneous (incomplete) pulses and adjust peak values if necessary

Ability to access a previous experiment, review a pulse with its metadata, markers and

Table 1. Identified tasks of the customized software program for data acquisition, storage

Ability to copy pulse or slope data to clipboard for further analysis

Distinguishing pulse data from non-pulse data

Allowing a fuzzy detection of the pulse peak

Viewing a representative pulse every 10 sec

Viewing decay rates (1/) in a second graph

associated notes

and analysis

Ability to pause and place a marker with a note

electrochemical potential Δψ (Ricci et al., 2004). Thus, induction of apoptosis is directly linked to mitochondrial dysfunction (Green and Kroemer, 2004).
