Cation-exchange high-performance liquid chromatography for variant hemoglobins and HbF/A2: What must hematopathologists know about methodology?

Table 1 Interpretation of various chromatogram regions and commonly encountered peaks in various high-performance liquid chromatography windows (retention time ranges obtained from Bio-Rad kit inserts)

Region/window	Retention time	Interpretation
Pre-integration region	< 1 min	Bilirubin, Hb H, Hb Barts, modified HbF
P1 peak	0.74	A minuscule peak usually found in specimens with increased HbF
F window	0.98-1.22	HbF, Hb Okayama
P2 window	1.28-1.50	Glycated HbA
P3 window	1.50-1.90	Aged samples, HbJ-Meerut, modified HbE
A window	1.90-3.10	HbA, Glycated HbS, intact Hb Koln
A2 window	3.30-3.90	HbE, Hb D-Iran, Hb Lepore, Hb G-Koushatta, Hb Zurich, Hb Korle Bu
D window	3.90-4.30	Hb D-Punjab, Hb G-Philadelphia
S window	4.30-4.70	HbS, Hb Q-Thailand, Hb Manitoba
C window	4.90-5.30	HbC, Hb Constant Spring, Hb Agenogi
Further unknown peaks	> 5.30	Hb Q-India