Transferrin isoforms

K9. Kleine, T. O., Damm, T, and Althaus, H., Quantification of /i-trace protein and detection of transferrin isoforms in mixtures of cerebrospinal fluid and blood serum as models of rhinorrhea and otorrhea diagnosis. Fresenius J. Anal. Cherrc 366,382-386 (2000). 

C. Whitfield JB. Transferrin isoform analysis for the diagnosis and management of hazardous or dependent drinking. CEn Chem 2002 48 2095-6. 

O. Martensson, A. HarUn, R. Brandt, K. Seppa and P. SiUanaukee, Transferrin isoform distribution gender and alcohol consumption. Alcohol. Clin. Exp. Res., 21, 1710-1715 (1997). 

E. Landberg, P. Pahlsson, A. Lundblad, A. Ametorp and J.-O. Jeppsson, Carbohydrate composition of serum transferrin isoforms from patients with high alcohol consumption, Biochem. Biophys. Res. Commun., 210, 267-274 (1995). 

A. Helander, G. Eriksson, H. Stibler and J.-O. Jeppsson, Interference of transferrin isoform types with carbohydrate-deficient transferrin quantification in the identification of alcohol abuse, Clin. Chem., 47, 1225-1233 (2001). 

J.M. Lacey, H.R. Bergen, M.J. Magera, S. Naylor and J.F. O Brien, Rapid determination of transferrin isoforms by immunoaffinity liquid chromatography and electrospray mass spectrometry, Clin. Chem., 47, 513-518 (2001). 

K. Yamashita, T. Ohkura, H. Ideo, K. Ohno and M. Kanai, Electrospray ionization-mass spectrometric analysis of serum transferrin isoforms in patients with carbohydrate-deficient glycoprotein syndrome, J. Biochem. (Tokyo), 114, 766-769 (1993). 

F.J. Legros, V. Nuyens, E. Minet, P. Emonts, K.Z. Boudjeltia, A. Courbe, J.L. Ruelle, J. Colicis, F. de L Escaille and J.P. Henry, Carbohydrate-deficient transferrin isoforms measured by capillary zone electrophoresis for detection of alcohol abuse, Clin. Chem., 48, 2177-2186 (2002). 

CDG-type I syndrome [264,266,267] is due to a deficiency in the oligosaccharidyltrans-ferase which transfers en bloc onto the nascent protein the oligosaccharide linked to dolichol diphosphate. Later, van Schaftingen and Jaeken [1995, FEES Lett. 377, 318-320] demonstrated that the syndrome was due in fact to a phosphomannomutase deficiency, an enzyme which provides the mannose-1-phosphate required for the initial steps of protein glycosylation. This leads to four transferrin isoforms non-glycosylated, glycosylated in Asn-413 or in Asn-611 and in both Asn-413 and 611. 

H. Henty and collaborators also showed alterations of the glycosylation of transferrin in patients with chronic alcohol abuse, producing the carbohydrate-deficient transferrin isoforms (Henry et al, 1999). These alterations were veiy similar to those present in patients with carbohydrate-deficient glycoprotein syndrome type 1 (CDGl) (Henty et al, 1997 Heyne... 

Arizaga RS, Blanco GE, Alvarez LG Montes-Bayon M, Sanz-Medel A. Detection of transferrin isoforms in human serum comparison of UV and ICP-MS detection after CZE and HPLC separations. Anal Bioanal Chem 2005 383 390-7. 

Legros, F.J. et al.. Carbohydrate-deficient transferrin isoforms measured by capillary zone electrophoresis for detection of alcohol abuse, Clin. Chem., 48, 2177, 2002. 

Kilar, F. and Hjerten, S., Separation of the hnman transferrin isoforms by carrier-free high-performance zone electrophoresis and isoelectric focnsing, J. Chromatogr. A, 480, 351,1989. 

Tagliaro, F. et al.. Optimized determination of carbohydrate-deficient transferrin isoforms in serum by capillary zone electrophoresis. Electrophoresis, 19, 3033, 1998. 

Several pure proteins have been purified and checked for their purity, microheterogeneity, or diagnostic significance by CE. In this case, coated capillaries are preferred for this separation. Many of the proteins can be analyzed by CE however, sensitive detectors such as fluorescence are necessary [68-70]. Examples of proteins studied by CE separately or in a profile are transferrin isoforms, which are important as markers of alcoholism [71], a-1 antitrypsin [72], recombinant human erythropoietin glycoforms that stimulates erythopiosis [73], plasminogen tissue activator [74], prions [75], urothelial carcinoma proteins in urine [76,77], and numerous urinary proteins [78]. 

In CZE, serum proteins can be separated also into 5-12 zones using different buffers such as Tris, borate, and tricine, with a pH of 8-11. Serum protein separation can be completed by CE in 2-10 min in contrast to 1-2 h for AG (Figure 2). The correlation coefficient between CE and AG for the separated bands is good. Transferrin isoforms, which are important as marker of alcoholism, have been separated by CZE. Some commercial instruments use multicapillaries of narrow diameter (25 pm) to increase the throughput of the analysis. The narrow capillaries produce better resolution than the wider capillaries with a shorter migration time. 

D-binding immunoaffinity solid phase is used in a batch mode to concentrate vitamin D analogs prior to reverse-phase LC separation and mass spectrometric quantification [16]. Alternatively, an immunoaffinity column can be used in the main separation step of analysis, as has been reported for analysis of transferrin isoforms [17], An important consideration in the use of immunoaffinity columns in the clinical laboratory is the increased cost of the method compared to more common solid phases and the fact that the life time of an immunoaffinity column is shorter than a standard reversed- or normal-phase column. 

Later, Busto et al. used synthesized Fe-labeled transferrin to determine individual transferrin isoforms in human serum samples for the species-specific mode. The stability of the prepared proteins was tested for 1 week and no iron exchange had occurred. They concluded that the results are in good agreement with other calibration methods, e.g. the species-unspecific method however, the species-spedfic mode can offer better precision. Hoppler et al. also synthesized and characterized Fe-labeled Phaseolus vulgaris ferritin for isotope dilution analysis. " ... 

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