CALCIUM IN SERUM BY ISE

Free ionic calcium may be determined using the technique of known addition after dilution of the serum sample.

Equipment Required

1. Ion analyzer (recommended) or pH meter with millivolt scale.
2. Calcium combination Ion selective electrode

Reagents

1. Calcium 10 mmol/l^-1 standard
2. 1 Molar KCl (ISAB)
3. Deionized water

Standard Preparation

Dissolve 14.702g CaCl₂.2H₂O (analar) in deionized water in a liter volumetric flask. Dilute to the mark with deionized water. This is 100 mmol/l^-1 Ca²⁺.

Place 10 ml of 100 mmol/l^- Ca²⁺ standard in a volumetric flask and dilute to the mark with deionized water. This is 10 mmol/l^- Ca²⁺ standard.

Prepare also a 1 mmol Ca²⁺ solution.

Sample Preparation

To 0.5 ml of serum in a 10 ml volumetric flask add 1 ml of 1 Molar KCl and dilute to the mark with deionized water.

Slope Determination

1. To 20 ml of 10 and 1 mmol/l^-1 Ca²⁺ solutions add 2 ml 1M KCl.
2. Measure the potential of the 1 mmol standard (mV₁).
3. Rinse the electrodes with deionized water and measure the potential of the 10 mmol/l^-1 standard (mV₂).

 

Electrode slope = mV₂ – mV₁ (mV/decade)

Method

1. Immerse the electrodes in 10 ml of the sample and record the mV potential (mV₁).
2. Add 1 ml of the 10 mmol/l^-1 Ca²⁺ standard - stir well and record the new potential (mV₂).

Calculation

If a pH/ion meter with a K add mode is being used the calculation of Ca²⁺ in the sample is performed automatically and displayed in direct concentration units of your choice. Multiply this value by 20 to account for the dilution factor.

If a pH meter with millivolt scale has been used operate the following calculation:

Where: Cu  =     concentration of the unknown

            Cs   =     concentration of the standard

Vs    =     volume of the standard (ml)

Vu    =     volume of the sample (ml)

ae    =     mV₂ – mV₁

S     =     electrode slope in mV