PubMed Health⌕ Search

Biomedical subjects

K Gerritse

Publications and source records attributed to K Gerritse.

11 recordsLinked to original sources
PubMed1996

CD40-CD40 ligand interactions in experimental allergic encephalomyelitis and multiple sclerosis.

We investigated the role of CD40-CD40 ligand (CD40L) interactions in multiple sclerosis (MS) and experimental allergic encephalomyelitis (EAE). Activated helper T cells expressing CD40L (gp39) surface protein were found in MS patient brain sections, but not in brain tissue sections of normal controls or patients with other neurological disease. CD40L-positive cells were co-localized with CD40-bearing cells in active lesions (perivascular infiltrates). Most of these CD40-bearing cells proved to be of the monocytic lineage (macrophages or microglial cells), and relatively few were B cells. To functionally evaluate CD40-CD40L interactions, EAE was elicited in mice by means of proteolipid-peptide immunization. Treatment with anti-CD40L monoclonal antibody completely prevented the development of disease. Furthermore, administration of anti-CD40L monoclonal antibody, even after disease onset, shortly before maximum disability score was reached led to dramatic disease reduction. The presence of helper T cells expressing CD40L in brain tissue of MS patients and EAE animals, together with the functional evidence provided by successful experimental prevention and therapy in an animal model, indicates that blockade of CD40-CD40L-mediated cellular interactions may be a method for interference in active MS.

Animals↗
PubMed1994

The involvement of specific anti myelin basic protein antibody-forming cells in multiple sclerosis immunopathology.

Irrespective of the large body of literature on the putative role of antibodies in the development of multiple sclerosis (MS), the detection of specific antibody-forming B cells (AFCs) in the central nervous system (CNS) tissues has not been described. In this study we show that autoantigen-specific AFCs can be found in CSN tissue sections of MS patients. Applying a newly developed myelin basic protein (MBP)-enzyme conjugate technique, we have detected MBP-specific AFCs in autopsy periventricular white matter and cerebellum tissue sections of MS patients. We demonstrated the presence of MBP-specific AFCs in CNS tissue sections in five out of 12 MS patients. No MBP-specific AFCs were detected in CNS tissue sections of 11 patients with other neurological diseases, such as Parkinson's and Alzheimer's disease, or in brain tissue sections of eight deceased persons without neurological diseases. In MS patients, anti-MBP AFCs were present in brain tissue sections both with and without plaques. The proportion of MBP-specific AFCs in some of the MS patient brain tissues reached over 50% of all AFCs. The high relative frequency of the anti-MBP AFCs and their localization in periventricular white matter and cerebellum of MS patients only, suggests that anti-MBP AFCs represent a cell population, which could play an important role in MS immunopathology.

Adult↗
PubMed1992

Immunological discrimination between the human apolipoprotein E2(Arg158----Cys) and E3 isoforms.

A specific anti-apoE2(Arg158----Cys) monoclonal antibody was raised by means of immunization of mice with a variant specific synthetic peptide. The peptide sequences used were homologous to apolipoprotein E of human and mouse. Consequently, the mouse immune system was tolerant to most of the selected sequences. Immunization with only one of selected peptides (amino acids 154-172) evoked an anti-peptide and anti-native protein response. Surprisingly, this peptide was predicted to have a low antigenicity index, in contrast to the other used peptides. The variant specific anti-peptide MAb that was generated with this sequence, recognizes apoE2(Arg158----Cys) and not apoE3. We here describe a sensitive, time saving, and simple immunoblot assay to detect apoE2(Arg158----Cys) in human sera without prior isoelectric focusing of serum proteins.

Amino Acid Sequence↗
PubMed1991

Conjugate formation in urea: coupling of insoluble peptides to alkaline phosphatase for ELISA and in situ detection of antibody-forming cells.

We report here a new method to produce synthetic peptide/alkaline phosphatase (AP) conjugates in the presence of urea. The method allows the use of peptides that are not soluble to a sufficient degree in aqueous buffers. The presence of 8 M urea during the construction of the synthetic peptide/AP conjugates does not influence enzyme activity nor the affinity of the anti-peptide antibodies for the conjugated peptide. We demonstrate that these synthetic peptide/AP conjugates can be used for detection of specific antipeptide antibody-forming cells (AFC) in vivo. This method for constructing enzyme conjugates with insoluble proteins or peptides suggest not only new possibilities for detection of specific AFC in vivo but also for applications in receptor-ligand studies, ELISA (enzyme-linked immunosorbent assay), and spot ELISA for detection of antibody-secreting cells in vitro.

Alkaline Phosphatase↗
PubMed1991

Detection of genetic variants of alpha 1-antitrypsin with site-specific monoclonal antibodies.

The serum protein alpha 1-antitrypsin (alpha 1-AT) serves as the major inhibitor of neutrophil elastase. The most common allele of the alpha 1-AT gene is designated as PiM. The Z mutation is a single-base substitution of the normal M allele, causing a Glu----Lys change at position 342 in the molecule. The ZZ phenotype is associated with a severe deficiency of alpha 1-AT, serum concentrations of the protein being 10% of normal. Individuals with an alpha 1-AT deficiency are at an increased risk of developing emphysema. To generate antibodies that specifically detect the 342 position in the context of the flanking sequences, we synthesized several peptides that included the 342 position for both the M and the Z variant. Immunization with variant-specific peptide-carrier conjugates elicited alpha 1-AT variant-specific responses, as determined in a direct enzyme-linked immunoassay. Monoclonal antibodies (MAbs) were selected with different specificity for the 342 region: MAbs F43 recognize only the alpha 1-AT sequence with 342Glu, i.e., all variant proteins that are non-Z, either from hetero- or homozygous individuals; MAbs F50 recognize only the sequence with 342Lys, i.e., all Z-variant proteins in ZZ or heterozygous individuals; MAbs F46 recognize alpha 1-AT with either 342Lys or 342Glu, all variant proteins with sequences as in the peptides used. Z homo- and heterozygotes were detected with our MAbs in a rapid and simple immunoblot assay. Other variants (M, S, and F) can also be assigned on the basis of the electrophoretic pattern. This sensitive detection method is very easy, rapid, and straightforward and provides a powerful tool for diagnosis of the alpha 1-AT deficiencies, allowing early treatment (augmentation of alpha 1-AT) and proper advice on lifestyle practices.

Animals↗
PubMed1990

An improved conjugation method for controlled covalent coupling of synthetic peptides to proteins using glutaraldehyde in a dialysis method.

Controlled and efficient conjugation of synthetic peptides to proteins, for use in immunization or in assay procedures, is a prerequisite for the immunological applications of synthetic peptides. This study describes a new method of conjugating synthetic peptides to proteins in such a way that no homopolymers of synthetic peptides or proteins occur. To achieve this, the protein is first activated with glutaraldehyde and subsequently excess glutaraldehyde is removed. Then coupling of the synthetic peptide to the activated protein occurs while subsequently the surplus reactive glutaraldehyde groups on the protein are blocked with lysine. Excess free peptide and lysine is then removed by dialysis. This improvement not only results in better defined conjugates when compared to classical glutaraldehyde coupling, but also in the consumption of smaller amounts of synthetic peptide during conjugate formation. When used for immunization we obtained similar and sometimes even better responses with the glutaraldehyde based conjugates than with succinimidyl (MBS) conjugates of the same peptides. The performance of the modified conjugates in ELISA procedures, immunization and immunocytochemistry suggests that they are superior to conjugates formed by classical glutaraldehyde coupling.

Amino Acid Sequence↗
PubMed1990

A novel carbodiimide coupling method for synthetic peptides. Enhanced anti-peptide antibody responses.

Coupling of peptides to immunogenic protein carriers is required for the generation of anti-peptide antibody responses. Carbodiimides are hetero bi-functional coupling reagents that are utilized for coupling reactions through carboxyl and amino groups. The procedures generally used for carbodiimide coupling of peptides and proteins result in conjugates which generate immunodominant antibody responses directed against the neodeterminants on the carrier protein. These determinants are induced by the reaction of carrier and/or peptide with the coupling agent. We have investigated the potential inhibiting effect of an imidazole intermediary on the formation of unwanted neodeterminants during carbodiimide coupling. The serum antibody responses elicited with the peptide-protein conjugates produced were evaluated in ELISA. We have modified and improved the coupling with a watersoluble carbodiimide (EDC) in such a way that a high response to the coupled peptide is obtained in association with negligible levels of anti-neodeterminant antibodies.

Amino Acid Sequence↗
PubMed1990

Specific immuno-detection of benzoate-para-hydroxylase with antibodies raised against synthetic peptides.

As a model system for the industrial use of fungal cells in the enzymatic conversion of chemicals, the parahydroxylation of benzoate was studied. To increase the amount of benzoate-para-hydroxylase (BPH, EC 1.14.13.12.) in the cell the gene coding for the enzyme (bphA) was cloned and expressed in Aspergillus niger. Detection of the enzymatic activity of the protein was not reproducible. It was decided to raise an antiserum for immuno-detection purposes. Sufficient benzoate-para-hydroxylase for immunization could not be obtained; therefore the synthetic-peptide strategy was used. We demonstrate that synthesis of antigenic determinants, can be useful in the production of highly specific reagents for the detection of proteins. The availability of monospecific polyclonal sera opens new possibilities in functional studies and purification of benzoate-para-hydroxylase.

Amino Acid Sequence↗
PubMed1990

Double immunocytochemical staining for in vivo detection of epitope specificity and isotype of antibody-forming cells against synthetic peptides homologous to human immunodeficiency virus-1.

Many infections evoke a strong humoral immune response. Some (e.g., HIV-1, EBV, CMV) also lead to disorders of the B-cell system. Data concerning cell dysfunction are largely derived from in vitro studies, which necessarily exclude all microenvironmental influences. The aim of this study was to develop a tool for the investigation of epitope specific humoral immune responses in vivo. Mice were immunized with one of two synthetic peptides, both 21 amino acids long and homologous to regions of the HIV-1 gp160. Cryostat sections of spleen and lymph nodes were incubated with the corresponding peptide coupled to alkaline phosphatase and simultaneously incubated with peroxidase-conjugated rabbit antisera specific for mouse immunoglobulin isotypes. We were able to show simultaneous detection of epitope specificity, isotype, and localization of antibody-forming cells and immune complexes in tissue sections. It should prove useful for in vivo investigation of the development of specific (e.g., anti-HIV-1) humoral immune response, the determination of B-cell specificity in lymph node infiltrates, and the role of immune complexes in lymph node pathology.

Alkaline Phosphatase↗
PubMedDate not supplied

Oral administration of TNP-Lactobacillus conjugates in mice: a model for evaluation of mucosal and systemic immune responses and memory formation elicited by transformed lactobacilli.

Safe live vector systems are being developed for oral delivery of antigens. A transformation system for indigenous Lactobacillus species of the gastrointestinal tract is described. Model systems were set up to evaluate immune responses. Orally administered trinitrophenylized (TNP) Lactobacillus were examined for their ability to induce immunological memory formation via determination of specific antibody titres in serum. We demonstrate a direct correlation between the level of systemic memory formation, as revealed by specific anti-TNP IgG serum antibodies, and the TNP substitution ratio of the Lactobacillus suspension used for oral priming. The specific IgG anti-TNP serum titres were comparable to or even higher than the titres of parental intraperitoneally primed animals. These results demonstrate the feasibility of using orally administered antigen-Lactobacillus as a future approach to vaccination.

Administration, Oral↗