Our Experience with Diagnostics of Congenital Disorders of Glycosylation

Abstract

The aim of this study is to report our 3years experience with the screening of congenital disorders of glycosylation. A common isoelectric focusing method with immunofixation was used for analysis of serum transferrin and α1-antitrypsin, apart from several other procedures. A group of about 1000 individuals, both healthy controls and patients, mostly with signs of a metabolic disease were examined. Here we present an overview of 1) hypoglycosylation findings, 2) distribution of protein variants, 3) misguiding rare Tf variants found in our set, and 4) association of some phenotypes with various diseases.

Microheterogeneity of some serum glycoproteins in neurodegenerative diseases

“…Analyses of serum Tf and aAT [33], ApoE [34] and ApoCIII [35] were performed by isoelectric focusing (IEF) with direct immunofixation and Coomassie or silver staining. For proper identification of protein variants, digestion with neuraminidase (from Clostridium perfringens, Sigma, St. Louis, USA) prior to IEF was used [33].…”

“…For proper identification of protein variants, digestion with neuraminidase (from Clostridium perfringens, Sigma, St. Louis, USA) prior to IEF was used [33]. Protein phenotypes detected by IEF correspond in fact to their genotypes, and this term is used hereafter.…”

Genetic variants of transferrin in the diagnosis of protein hypoglycosylation

“…IEF was carried out (Albahri et al 2004), using Immobiline DryPlate gels of pH 4-7, or alternatively Ampholine PAGplate gels of pH 3.5-9.5 (Amersham Pharmacia Biotech), with detection by immunofixation and Coomassie Blue staining. Digestion with neuraminidase (No.…”

Transferrin D protein variants in the diagnosis of congenital disorders of glycosylation (CDG)

“…A common IEF method used for Tf (and possibly also for a 1 -antitrypsin, aAT) analysis with direct immunofixation was described earlier (7). In short, a Pharmacia Multiphor apparatus cooled to 121C, Immobiline DryPlate gels pH 4-7 and 5-6 or CleanGel IEF with Ampholyte pH 3-10 (GE Healthcare, Uppsala, Sweden), PhastGel Blue R (Coomassie brilliant blue R-350, Amersham Biosciences, Uppsala, Sweden) staining and densitometric evaluation were used for the CDG screening.…”

“…Horizontally: isoforms of Tf C1 and Tf D2 and Tf D-x (unknown variant) are indicated; 0-6, a-, mono-, di-, tri-, tetra-, penta-, and hexa-sialo-Tf. For Tf analysis, all samples were iron-saturated with Fe-citrate and diluted 1 in 5, as described in (7). The IEF procedure was further amended by pre-treating the sample with neuraminidase (from Cl.…”