Method F-003M Calcium in Biological Fluids

The mixture is then centrifuged and the supernatant fluid decanted off to be used for sodium and potassium determination if required.

The precipitate is dissolved in perchloric acid and then aspirated into the calibrated Sherwood Flame photometer.

Equipment Required;

1. Sherwood Model 410 Flame Photometer
2. Balance weighing to +-0.0005g

Reagents Preparation:

1. Dissolve 12.6g (C00H)2 2H₂0 to 1 litre distilled water in a volumetric flask. This is 0.1 M oxalic acid.
2. Dissolve 12.2g (C00NH₄)₂ H₂O to 1 litre of distilled water in a volumetric flask. This is 0.1 M ammonium oxalate.
3. To 5ml of 0.1 M oxalic acid in a 100ml volumetric flask add 0.1 M ammonium oxalate up to the mark.

0.05M Perchloric acid.

Standard Preparation:

1. Pipette exactly 100 mls Ca standard (Sherwood part 00156623) into 1 litre volumetric flask make to mark with 0.05 m perchloric acid reagent. This is the calcium 100mg/1 standard.
2. A range of standards is now prepared by successive dilution of the 100mg/l standard with 0.05M perchloric acid.

Method:

1. Pipette 2mls of serum into a rounded centrifuge tube and add 3mls of Ammonium oxalate/oxalic acid mixture.
2. Mix and stand for 30 minutes.
3. Centrifuge well and decant the supernatant fluid and drain the sample.
4. Add 4ml of 0.05M perchloric acid to the pellet, vortex or shake vigoriously to dissolve the precipitate fully. The sample is now ready for analysis.
5. Treat 2ml of the calcium 100mg/l standard in the same way as the sample, ie. stages 1 -4, to obtain a control.
6. Select the calcium filter on the flame photometer.
7. Aspirate the top calcium standard into the Sherwood Flame photometer and adjust reading to 100
8. Without touching the controls aspirate the other standards and check linearity
9. Aspirate the sample solution into the flame photometer and read the calcium concentration directly from the Display.
10. Aspirate the control into the flame photometer and record reading.

Calculation