Labile HbA1c migrates in the #C fraction together with modified hemoglobin (such as carbamylated hemoglobin, acetaldehyde hemoglobin, and acetylated hemoglobin) when HbA1c is measured by Arkray's high-performance liquid chromatography (HPLC). It is assumed that most of the labile glycation products of variant hemoglobin do not migrate in #C fraction; in addition, a part of the stable glycation products of variant hemoglobin migrates in #C fraction. We hypothesized that subjects with variant hemoglobin are likely to show abnormally low or high values of #C fraction. In this study, we investigated this hypothesis. Twenty-one non-diabetic subjects with nine types of variant hemoglobin, and 103 non-diabetic subjects without variant hemoglobin were used. HbA1c and #C fraction were measured by Arkray's HPLC (HA-8180) using standard mode. The values of #C fraction in the control group were 1.75 ± 0.15% (range: 1.5-2.1%). The variant hemoglobin group reported #C fraction values of ≤1.3% in twelve subjects, ≥2.3% in five subjects, and within the reference range (1.4-2.2%) in three subjects. When the cutoff values of #C fraction were set at ≤1.3% and ≥2.3%, sensitivity and specificity were 86% and 100%, respectively. Most non-diabetic subjects with variant hemoglobin showed abnormal values of #C fraction. Measurement of #C fraction is a useful screening test for variant hemoglobin in non-diabetic subjects.