CZE of human alpha-1-acid glycoprotein for qualitative and quantitative comparison of samples from different pathological conditions

Abstract

Alpha-1-acid glycoprotein (AGP) presents different forms, which may arise from dif- ferences in the amino acid sequence and/or in the glycosidic part of the protein. Changes in forms of AGP have been described in literature as a possible tumor marker. While most previous works have approached the study of glycopeptides and/or gly- cans obtained after fragmentation of the protein, in this work, a CZE method is devel- oped to separate up to eleven peaks of intact forms of AGP. A computer program developed in our laboratory is used to select the migration parameters that make possible an accurate assignment of AGP peaks. Electropherograms of AGP samples purified from sera of cancer patients and healthy donors are qualitatively and quanti- tatively compared. Percentages of correct assignment of AGP peaks close to 100% are achieved by using either the migration time of each peak relative to that of the EOF marker or the effective electrophoretic mobility of the peaks. The computer program permits to select, among different hypotheses for peak allotment, that one providing the highest accuracy of assignment. In this way, some peaks with different charge-to-mass ratio and a different distribution of area percentage of AGP forms are observed when comparing samples from sick and healthy individuals. Thus, a method that permits to compare AGP forms existing in sera of individuals with different pathophysiological situations has been developed. A potential for using AGP forms analyzed by CZE as a disease marker and for using this technique for screening purposes is envisaged.The Authors greatly acknowledge Prof. Kremmer for the kind gift of AGP samples. This work has been supported by Spanish CICYT, Plan I1D1I (Project TIC2003-01906), by Foundation Ramon Areces, and by Ministry of Education and Science (Joint Action with Hungary HH04-33). Izaskun Lacunza acknowledges CSIC for a predoctoral fellowship.Peer reviewe