Table 1 Summary of the methodology from each of the four ICD protocols used in this experiment

ICD-1

Mix 100 µl of serum or plasma with 100 µl 0.1 M disodium EDTA (pH 7.5) (1:1 ratio) and incubate at 100 °C for 5 min, followed by centrifugation at 16000×g for 5 min

Weil et al. [8]

ICD-2

Mix 600 μl serum or plasma sample with 200 μl 0.1 M disodium EDTA (pH 7.5) (ratio 1:3) and incubate at 104 °C for 10 min. Then, centrifugate at 16000×g for 5 min

Swartzentruber et al. [12]

ICD-3

Mix 100 µl of serum with 100 µl of 7.5% (w/v) TCA (1:1 ratio) to achieve pH = 1. Incubate at room temperature for 20 min followed by centrifugation at 16000×g for 5 min. Approximately 80% (170 μl) of the total starting volume of serum or plasma and TCA is recovered. Mix 150 μl aliquot of the centrifugated sample + 30 μl 1 M Trizma buffer (in a volume equal to 20% of the volume of the recovered supernatant) and then invert several times to mix, thus returning the sample to a neutral pH (pH = 7–8)

Starkey et al. [21]

ICD-4

Mix 100 μl serum or plasma sample + 300 μl SDS 7 mM; DTPA, 1.5 mM (pH 7.2) solution (1:3 ratio) vortex and incubate at 95–98 °C for 4 min

Steindl et al. [10]