α1-Antitrypsin in serum determined by capillary isoelectric focusing

Abstract

A capillary isoelectric focusing (CIEF) method using bare fused-silica capillaries filled with polyethylene oxide (PEO) and carrier ampholyte solutions in the pH 3.5-5.0 range has been developed for the identification of alpha1-antitrypsin (alpha1AT) phenotypes in human serum. This novel procedure was routinely applied to the study of serum samples of five controls whose alpha1AT phenotype was previously identified and of twelve subjects whose alpha1AT phenotype was unknown. The results obtained allowed us to confirm or identify the alpha1AT phenotype in all sera tested. This procedure seems particularly suitable for identification of alpha1AT variants associated with diseases of clinical relevance.

SP-A binds alpha1-antitrypsin in vitro and reduces the association rate constant for neutrophil elastase

“…Wild-type α 1 -antitryspin (M α 1 -AT) was either from ART or purified from human serum by covalent chromatography. Capillary isoelectric focusing (CIEF) with bare fused-silica capillaries filled with polyethylene oxide and carrier ampholyte solutions in the pH 3.5–5.0 range [ 14 ] was applied to confirm the presence of the common M α 1 -AT variant. Z α 1 -AT variant was purified by covalent chromatography from subjects identified within the Italian screening program for α 1 -AT deficiency [ 15 ].…”

Determination of the isoelectric point of proteins by capillary isoelectric focusing

“…I let you draw your conclusions, but it would seem that never mind the second and third decimal digits, even the first decimal appears to be off the target! More modestly, Lupi et al [71], in an extensive investigation on the polymorphism of ␣ 1 -antitrypsin in sera, have given the pI values of the various isoforms with only two decimal digits. Also, Cifuentes et al [72], even though they reported exquisite resolution of seven isoforms of recombinant human erythropoietin (see their Fig.…”

Capillary electrophoresis of proteins 1999-2001

“…A dynamic CIEF simulator has been developed to model separation of up to 150 components [124]. CIEF has been used to analyze cellulase composition [128], microbial proteomes [129,130], a 1 -antitrypsin phenotypes in serum [131], proteins in human cerebrospinal fluid (CSF) [121], noncovalent protein complexes [132], isoenzymes of human alcohol dehydrogenase [133], FQlabeled green fluorescent protein [134], yeast cytosol proteins [135], HIV envelope glycoprotein [44], and protein heterogeneity caused by labeling with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate [29].…”