Blood Gas Analysis

Describe the methods of measurement of oxygen and carbon dioxide tension in blood and blood pH

Blood gas machines directly measure three variables and calculate the remainder. Measured variables are:

• PO₂
• CO₂
• pH

Calculated variables include:

• Bicarbonate
  Using the pH, CO₂ and the Henderson-Hasselbalch equation.
• Base Excess
  Calculated using the Henderson-Hasselbalch and Siggaard-Anderson equation. Can be expressed in two ways:
  • Base Excess
    The amount of alkali that must be added to the sample to return it to a normal pH, at a temperature of 37°C and a PaCO₂ of 40mmHg.
  • Standardised Base Excess
    As base excess, but calculated for blood with a Hb concentration of 50g.L^-1. This is thought to better represent the ECF as a whole.

Oxygen Tension

Oxygen tension is measured with a Clarke electrode. This consists of:

• A chamber for the blood sample
• A chamber containing a potassium chloride solution, which:
  • Is separated from the blood chamber by an oxygen-permeable membrane This prevents blood being in direct contact with the cathode, which would lead to protein deposition on the cathode and incorrect measurement.
  • Contains a platinum cathode
  • Contains a silver/silver Chloride anode
• A battery applying 0.6V across the electrodes

Method

• A voltage of 0.6V is applied across the electrodes, causing the silver to react with chloride in the solution to produce electrons:
  $Ag + Cl^- \Rightarrow AgCl + e^-$
  • This potential difference is required to start the reaction
  • 0.6V is chosen because it is enough to start the reaction but will have minimal effect on measured current flow
• At the cathode, oxygen combines with electrons and water to produce hydroxyl ions: $O_2 + 4e^- + 2H_2O \Rightarrow 4OH^-$
• For each oxygen molecule present at the cathode, four electrons can be consumed
• Increasing the oxygen available at the cathode increases the number of electrons consumed, and therefore increases current flow
  • Oxygen will move from the sample chamber to the measuring chamber according to its partial pressure
• Measured current flow is therefore proportional to oxygen tension in blood

Calibration, Limitations, and Accuracy

• Calibration is performed with standard gas mixtures
  Requires regular two-point calibration.
• Cathode must be kept clean from protein and not damaged
• Cathode must be kept at 37°C
• May read falsely high with halothane

pH Measurement

pH is a measure of the hydrogen ion concentration¹ in solution, and is defined as the negative logarithm to the base 10 of the [H⁺]:

• $pH = -log_{10}[H^+]$
• A pH of 7.4 is a [H⁺] of 40nmol.L^-1 at 37°C
  • A change in a pH unit of 1 is equivalent to a 10-fold change in the [H⁺]
  • A change in pH of 0.3 is equal to doubling or halving the [H⁺]

The pH electrode consists of:

• A chamber for the blood sample
• A measuring chamber, separated from the sample by H⁺-permeable glass, which contains:
  • A buffer solution
  • A silver/silver chloride measuring electrode
• A reference chamber, also separated from the chamber by H⁺-permeable glass, which contains:
  • A KCl solution
    Has no buffering properties.
  • A mercury/mercury chloride reference electrode

Method

• Relies on the principle that two solutions with different H⁺ activities will develop a potential difference between them (proportional to the concentration gradient)
• H⁺ passes through the glass along a concentration gradient:
  • A variable potential difference is generated in the measuring chamber, as H⁺ ions are buffered and the concentration gradient is maintained
  • A constant potential difference is generated in the reference chamber, as there is no buffer of H⁺ ions in the KCl solution
• Once H⁺ has equilibrated between blood and the KCl solution, the potential difference between the measuring and reference electrodes is proportional to the H⁺ concentration in blood

Calibration, Limitations, and Accuracy

• Calibration is performed with two phosphate buffer solutions containing two different (known) [H⁺]
• Must be kept at 37°C
  Hypothermia increases solubility of CO₂ and therefore lowers PaCO₂
  A reduced partial pressure of CO₂ is required to keep the same number of molecules dissolved (as per Henry's Law)
  • Therefore, as blood cools its pH will increase
• Electrodes must be kept clean from protein and not damaged

Carbon Dioxide Tension

Carbon dioxide tension is measured with a Severinghaus electrode, which is based on the pH electrode, as PaCO2 is related to [H⁺]. The Severinghaus electrode consists of:

• A chamber for the blood sample, separated from the bicarbonate chamber by a CO₂ permeable membrane
• A chamber containing bicarbonate solution in a nylon mesh, and separated from both the measuring and reference chambers by H⁺-permeable glass
• A measuring chamber containing:
  • A buffer solution
  • A silver/silver chloride measuring electrode
• A reference chamber containing:
  • A KCl solution
  • A mercury/mercury chloride reference electrode

Method

• CO₂ diffuses from blood into the bicarbonate chamber
• CO₂ reacts with water in the bicarbonate chamber to produce H⁺ ions
• From here, the process is identical to the pH electrode, except bicarbonate takes the place of blood:
  • H⁺ ions diffuse into the reference chamber until the H⁺ ion concentration has equilibrated
  • H⁺ ions continually diffuse into the measuring chamber (as they are buffered)
    • This establishes a constant pH gradient
      This gradient is proportional the H⁺ ion concentration in the bicarbonate chamber, which is proportional to the CO₂ content of blood.

Calibration, Limitations, and Accuracy

• Calibration is performed with solutions of known CO₂ concentration
• Must be kept at 37°C
  Hypothermia decreases solubility of CO₂ and therefore decreases pH
• Electrodes must be kept clean from protein and not damaged
• Slow response time relative to pH electrode due to time taken for CO₂ to diffuse and react
  This can be accelerated with carbonic anhydrase

Footnotes

¹. Technically pH is defined as the activity of H⁺ in a solution. Clinically, activity is identical to concentration, so in medicine these definitions are functionally the same. ↩

References

1. Leslie RA, Johnson EK, Goodwin APL. Dr Podcast Scripts for the Primary FRCA. Cambridge University Press. 2011.
2. (FRCA - Measurement of pO2, pCO2, pH, pulse oximetry and capnography)[http://www.frca.co.uk/article.aspx?articleid=100389]
3. Aston D, Rivers A, Dharmadasa A. Equipment in Anaesthesia and Intensive Care: A complete guide for the FRCA. Scion Publishing Ltd. 2014.