| 1. |
- Lönnberg, Maria, 1953-
(författare)
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Membrane-Assisted Isoform ImmunoAssay : Separation and determination of protein isoforms
- 2002
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Proteins exist in a variety of isoforms with minor differences, mostly due to their glycosylation patterns, which can modulate their biological functions. It seems to be of clinical relevance to measure the isoform-distribution.Thesis describes a novel technology named Membrane-Assisted Isoform ImmunoAssay (MAIIA). This technique allows rapid (< 15 min.) isoform determination. It is based on a chromatographic separation combined with immunoassay detection. These steps are performed along a thin, disposable micro-porous chip in which capillary forces maintain the flow. By using anion-exchange as a chromatographic principle the technology has been utilized for the determination of transferrin isoforms in ten minutes. In one variant (the one-dimensional), selected isoforms (carbohydrate-deficient transferrin) are quantified. In a more elaborate variant (the two-dimensional) it was possible to determine the entire isoform profile of transferrin. Isoforms differing by only 0.1 pH unit in isoelectric point could be distinguished.The chromatography along the microporous bed of nitrocellulose showed very good separation performance with plate heights of 10-20 µm and only minor flow rate variations between individual devices. The quantitative determination of antibody-captured molecules was performed by using antibodies labelled with carbon black particles. Combined with a detection procedure by means of a flatbed scanner, a highly sensitive and specific immunoassay with a detection limit of 0.13 pM was obtained upon using IgE as a model analyte.This technology can thus be used to rapidly distinguish proteins with minor structure differences and specifically determine protein isoforms in complex environments, e.g., blood, down in the pM (10-12 M) concentration range.
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| 2. |
- Staff, Caroline, et al.
(författare)
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Induction of IgM, IgA and IgE Antibodies in Colorectal Cancer Patients Vaccinated with a Recombinant CEA Protein
- 2012
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Ingår i: Journal of Clinical Immunology. - : Springer Science and Business Media LLC. - 0271-9142 .- 1573-2592. ; 32:4, s. 855-865
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Tidskriftsartikel (refereegranskat)abstract
- Previous clinical studies have indicated that natural IgM antibodies have the ability to induce apoptosis of tumor cells but IgE and IgA may also mediate tumor cell killing (in addition to IgG). The aim of the study was to analyse induction of IgM, IgA and IgE antibodies in patients vaccinated with the tumor associated antigen CEA. Twenty-four resected CRC patients without macroscopic disease were immunized seven times with CEA +/- GM-CSF. Four different dose schedules were used over a 12-month period. IgM, IgA and IgE antibody responses against recombinant CEA were determined by ELISA. Patients were monitored immunologically for 36 months and clinically for 147 months. GM-CSF significantly augmented the anti-CEA response for all three antibody classes. Low dose of CEA tended to induce a higher IgM, IgA or IgE anti-CEA antibody response than higher. Anti-CEA IgA antibodies could lyse CEA positive tumor cells in antibody dependent cellular cytotoxicity (ADCC) as well as in complement dependent cytotoxicity (CDC). A significant correlation between survival and high IgA anti-CEA titers was noted (p = 0.02) irrespective of GM-CSF treatment. The observation that IgA anti-CEA antibodies were cytotoxic and associated with improved survival might indicate that also these antibodies may exert a clinical anti-tumor effect.
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| 3. |
- Ullenhag, Gustav J, et al.
(författare)
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Induction of IgG subclass responses in colorectal carcinoma patients vaccinated with recombinant carcinoembryonic antigen.
- 2002
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Ingår i: Cancer Research. - 0008-5472 .- 1538-7445. ; 62, s. 1364-
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Tidskriftsartikel (refereegranskat)abstract
- There is scanty information on the IgG subclass response after vaccination against cancer antigens. The induction and development of the IgG subclass responses in 18 colorectal carcinoma patients vaccinated s.c. seven times with recombinant human carcinoembryonic antigen (rhCEA) over a 12-month period were analyzed by ELISA. The patients were followed for 3 years. Four rhCEA doses were used, and half of the patients also received granulocyte macrophage-colony stimulating factor (GM-CSF) as an adjuvant. Anti-rhCEA-specific IgG1 and IgG4 responses and, to a lesser degree, IgG2 responses were markedly enhanced by concomitant GM-CSF administration, whereas the antigen dose was of minor importance. Almost no IgG3 response was observed. A significant antibody response was noted within the first weeks for IgG1 and IgG2 but noted several months later for IgG4. The responses gradually increased by repeated immunizations and peaked around 12 months for IgG1 and a few months later for IgG2 and IgG4. A sustained but reduced response was noted for these three subclasses at 24 and 36 months. Interestingly, there was a gradual shift from a predominant IgG1 response at 6 months to an IgG4 response at 15 months. No significant change in total concentrations of the four IgG subclasses was observed comparing prevaccination concentrations with concentrations at 12 months, indicating an antigen-specific effect of GM-CSF administration on the anti-rhCEA response. The clinical significance of the individual IgG subclass antibodies for tumor response is not clear and requires additional studies.
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| 4. |
- Wingren, Christer, et al.
(författare)
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Antigen-binding sites dominate the surface properties of IgG antibodies
- 1995
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Ingår i: Molecular Immunology. - 1872-9142 .- 0161-5890. ; 32:11, s. 819-827
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Tidskriftsartikel (refereegranskat)abstract
- A new technique, liquid-liquid partition chromatography in an aqueous polyethylene glycol-dextran two-phase system, was used to detect differences in surface properties of antibodies with different antigen-binding sites. Employing well-characterized monoclonal IgG antibodies and Fab and Fc fragments thereof as well as chimeric IgG antibodies we found a remarkable relationship between structure of the antibody combining site and chromatographic behaviour. The surface properties of the IgG antibodies were dominated by those of its antigen-binding regions. In addition, our results indicated that the constant parts of the IgGs form similar scaffoldings, on to which CDRs of variable shapes and sizes are interspaced and constitute the major dominant differences in exposed surface properties.
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