Comparison of α‐1‐acid glycoprotein isoforms from healthy and cancer patients by capillary IEF

Lacunza, Izaskun; Kremmer, T.; Diez-Masa, Jose Carlos; Sanz, J.; Frutos, Mercedes de

doi:10.1002/elps.200600700

Cited by 17 publications

(12 citation statements)

References 17 publications

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“…Figure 4 shows a plot of current vs. time for an open vs. packed capillary as focusing proceeds, with a fixed electric field of 200 V/cm. The current in the open and packed capillaries is on μA level, which is comparable to the value in regular cIEF as reported in the literature 19, 29. The current is initially three times higher for the open capillary, but the current decreases earlier.…”

Section: Resultssupporting

confidence: 86%

Field-Free Remobilization of Proteins after Isoelectric Focusing in Packed Capillaries

2010

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Pressure-driven remobilization without an applied electric field is shown to be possible with capillary isoelectric focusing by using packed capillaries. The capillary dimensions are 100 μm i.d. and 2 cm in length, and the packing is made of 0.9 μm nonporous silica particles that are chemically modified with a brush layer of polyacrylamide. Both reversible and irreversible adsorption are shown to be negligible. The packed capillaries eliminate the problem of unwanted hydrodynamic flow between reservoirs. Three proteins are focused: trypsin inhibitor, carbonic anhydrase II and myoglobin. The time required for focusing in the packed capillaries is increased by only a factor of two compared to the open capillary, giving complete focusing in less than 15 min at 200 V/cm. The packed capillaries allow the use of higher electric fields, with resolution continually increasing up to at least 1500 V/cm. The packing obstructs diffusional broadening after the field is turned off: for trypsin inhibitor, D=6.1(±0.3)×10-8 cm2/s for the packed capillary vs. D=28.8(±0.3)×10-8 cm2/s for the open capillary. The broadening contributed by the packing during remobilization is from eddy diffusion, and it is described by its plate height, H, which is the variance per unit length: H=σ2/L=0.64 μm. This limits the resolution to 0.1 pH unit for the 2-cm capillary having a pH range of 3-10, giving a theoretical peak capacity of 47.

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Section: Resultssupporting

confidence: 86%

Field-Free Remobilization of Proteins after Isoelectric Focusing in Packed Capillaries

2010

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“…In previous work, fucosylated haptoglobin was suggested as a biomarker for early detection of pancreatic cancer[27]. Also the glycoforms of alpha-1-acid glycoprotein have been found to vary in cancer patients compared to the healthy controls[28]. These biomarkers can be used to improve the confidence of the diagnosis through identification of disease-related glycan structures by various separation and mass spectrometry techniques[29-32].…”

Section: Introductionmentioning

confidence: 99%

Pancreatic Cancer Serum Detection Using a Lectin/Glyco-Antibody Array Method

et al. 2008

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Pancreatic cancer is a formidable disease and early detection biomarkers are needed to make inroads into improving the outcomes in these patients. In this work lectin antibody microarrays were utilized to detect unique glycosylation patterns of proteins from serum. Antibodies to four potential glycoprotein markers that were found in previous studies were printed on nitrocellulose coated glass slides and these microarrays were hybridized against patient serum to extract the target glycoproteins. Lectins were then used to detect different glycan structural units on the captured glycoproteins in a sandwich assay format. The biotinylated lectins used to assess differential glycosylation patterns were Aleuria aurentia lectin (AAL), Sambucus nigra bark lectin (SNA), Maackia amurensis lectin II (MAL), Lens culinaris agglutinin (LCA), and Concanavalin A (ConA). Captured glycoproteins were evaluated on the microarray in situ by on-plate digestion and direct analysis using MALDI QIT-TOF mass spectroscopy. Analysis was performed using serum from 89 normal controls, 35 chronic pancreatitis samples, 37 diabetic samples and 22 pancreatic cancer samples. We found that this method had excellent reproducibility as measured by the signal deviation of control blocks as onslide standard and 41 pairs of pure technical replicates. It was possible to discriminate cancer from the other disease groups and normal samples with high sensitivity and specificity where the response of Alpha-1-β glycoprotein to lectin SNA increased by 69% in the cancer sample compared to the other non-cancer groups (95% confidence interval 53% to 86%). These data suggest that differential glycosylation patterns detected on high throughput lectin microarrays are a promising biomarker approach for the early detection of pancreatic cancer.

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“…These glycoforms are absent in healthy controls as well as in treated ENL patients. Lacunza et al (2007) showed an increase in the acidic glycoforms of AGP by capillary IEF in ovarian cancer and lymphoma patients when compared with healthy controls. The tri-and tetra-antennary oligosaccharides included fucose residues that form the sialyl Lewis x (sLe x ) structures (Ceciliani & Pocacqua, 2007).…”

Section: Glycosylation Pattern Of Agp In Leprosymentioning

confidence: 87%

α 1-Acid glycoprotein as a putative biomarkerfor monitoring the development of the type II reactional stage of leprosy

Gupta

Dharmalingam

2010

Journal of Medical Microbiology

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Leprosy, a spectral disease manifested on the basis of host immune responses, is complicated by its reactional stages, namely type I reversal reaction (RR) and type II erythema nodosum leprosum (ENL). These reactional stages are characterized by uncontrolled and aberrant immune responses. Biomarkers for reactional stages would aid in early diagnosis, efficient treatment, prevention of neurological complications and prediction of predisposition to reactional stages. In this study, comparative analysis of the serum proteome of leprosy patients by two-dimensional electrophoresis (2DE) followed by mass spectrometry showed differential expression of acute-phase protein alpha (1)-acid glycoprotein (AGP; also known as orosomucoid). AGP levels in untreated ENL cases were significantly higher than in lepromatous leprosy (LL; a non-reactional disease stage) (P=0.0126), RR (P=0.0176) and healthy controls (P=0.0030). These data were confirmed using ELISA. The levels of AGP decreased to normal levels after treatment with multidrug therapy and thalidomide (P =0.0167). In a follow-up study, AGP levels, which were high in the untreated ENL stage, decreased significantly at 5 days ( P=0.0084) and 21 days (P=0.0027) post-treatment. A stage-dependent increase in AGP in an LL patient who progressed into the ENL stage was also shown. Glycosylation analysis by 2DE showed differential expression of acidic glycoforms of AGP in untreated ENL cases. Changes in AGP concentration and differential expression of isoforms correlated with the inflammatory condition in ENL and also with the treatment regimen. Thus, initial validation of AGP as an ENL-specific biomarker and treatment indicator was shown in this study.

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Comparison of α‐1‐acid glycoprotein isoforms from healthy and cancer patients by capillary IEF

Cited by 17 publications

References 17 publications

Field-Free Remobilization of Proteins after Isoelectric Focusing in Packed Capillaries

Field-Free Remobilization of Proteins after Isoelectric Focusing in Packed Capillaries

Pancreatic Cancer Serum Detection Using a Lectin/Glyco-Antibody Array Method

α 1-Acid glycoprotein as a putative biomarkerfor monitoring the development of the type II reactional stage of leprosy

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