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Biomedical subjects

L J Karr

Publications and source records attributed to L J Karr.

7 recordsLinked to original sources
PubMed1988

Cell separation by immunoaffinity partitioning with polyethylene glycol-modified protein A in aqueous polymer two-phase systems.

Previous work has shown that polyethylene glycol (PEG)-bound antibodies can be used as affinity ligands in PEG-dextran two-phase systems to provide selective partitioning of cells to the PEG-rich phase. In the present work we show that immunoaffinity partitioning can be simplified by use of PEG-modified Protein A which complexes with unmodified antibody and cells and shifts their partitioning into the PEG-rich phase, thus eliminating the need to prepare a PEG-modified antibody for each cell type. In addition, we provide a more rigorous test of the original technique with PEG-bound antibodies by showing that it is effective at shifting the partitioning of either cell type of a mixture of two cell populations.

Animals↗
PubMed1987

Phase partitioning in space and on earth.

In aqueous solution at low concentrations, the neutral polymers dextran and poly(ethylene glycol) (PEG) rapidly form a two-phase system consisting of a PEG-rich phase floating on top of a dextran-rich phase. Biological particles and macromolecules tend to partition differentially between the phases and the liquid-liquid phase interface in these systems. Bioparticle partitioning has been shown to be related to physiologically important surface properties such as membrane charge or lipid composition. Affinity partitioning into the PEG-rich phase can be accomplished by coupling PEG to a ligand having affinity for specific cells or macromolecules. Subpopulations can be identified or separated using multi-step countercurrent distribution (CCD). Incomplete understanding of the influence of gravity on the efficiency and quality of the impressive separations achievable by partitioning, and appreciation for the versatility of this efficient technique, have led to its study for low-gravity biomaterials processing. On Earth, two-phase systems rapidly demix because of density differences between the phases. In low-gravity, demixing has been shown to occur primarily by coalescence. Polymer surface coatings, developed to control localization of demixed phases in low-g, have been found to control electroosmosis which adversely affects electrophoretic separation processes on Earth and in space. In addition PEG-derivatized antibodies have been synthesized for use in immunoaffinity cell partitioning.

Dextrans↗
PubMed1986

Immuno-affinity partition of cells in aqueous polymer two-phase systems.

Poly(ethylene glycol) (PEG) was covalently coupled to IgG antibody preparations directed against human red blood cells. This modification reduces the tendency of the antibody to agglutinate cells and increases its affinity for the upper phase in dextran-PEG aqueous two-phase systems. These effects are related to the molecular weight of the PEG used for modification and to the number of PEG molecules attached to the antibody. Exposure of human red blood cells to PEG-modified antibody causes a substantial and specific increase in cell partition into the PEG-rich phase in a number of PEG-dextran aqueous two-phase systems. Pertinent phase-system parameters were examined. Following a single incubation with PEG-derivatized antibody, a mixture of sheep and human red blood cells was completely separated in 100 min by a 30-transfer countercurrent extraction using a two phase system which normally offers little resolution.

Animals↗
PubMedDate not supplied

Otitis media in the young infant: an IgE-mediated disease?

IgE antibody directed against noncapsular antigens of mechanically disrupted Streptococcus pneumoniae, serotype 3 rough, was demonstrated in middle ear effusions (MEE) and serum of infants with and without prior evidence of pneumococcal otitis media with effusion (OME). The techniques employed included radioimmunoassay (RIA), passive skin testing, Prausnitz-Küstner (P/K), and enzyme-linked immunospecific assay (ELISA). Adsorption of MEE with ultrasonically disrupted crude pneumococcal antigen (CPA-U) resulted in a reduction of total IgE counts per minute and suggested bacteria-specific IgE antibody ranging from approximately 22 to 92% of the total IgE. The biological activity of the IgE antibody was confirmed by challenging skin passively sensitized with MEE IgE and CPA-U. Areas of induration appeared 20 minutes after challenge and continued to increase in size until 90 minutes. An ELISA procedure was developed as a tool to determine the nature of the antigen(s) and to determine the class(es) of antibody other than IgE. It appeared that CPA-U possesses free amino groups and that it can withstand the rigors of autoclaving. An analysis of 45 cord bloods revealed that high levels of IgG CPA-U antibody occur in this type of sample and that no correlation exists between the IgE and IgG levels. The mean IgG:IgE ratio, optical density at 420 nm (OD 420), for cord bloods was 2.49. In contrast, serum samples from nine infants without pneumococcal otitis media and from 14 infants with pneumococcal otitis media had lower levels of IgG antibody. There was no significant relationship between IgG and IgE OD 420 in infants who never had an episode of pneumococcal otitis media and the mean IgG:IgE was 1.88, whereas the ratio for those infants with pneumococcal otitis media was 1.56. In addition, there was a significant correlation between IgG and IgE levels in this latter group. The results suggest that it may be important to monitor the levels of at least these two classes of antibody to enhance our understanding of the pathogenesis and recovery from otitis media.

Antibodies, Bacterial↗
PubMedDate not supplied

Immunologic response to pneumococcal polysaccharide vaccine in infants.

The serum antibody response to purified pneumococcal capsular polysaccharides (PCP) was detrmined in four groups of infants ranging in age from 3 to 24 months. Group 1 consisted of eight infants immunized with an octavalent vaccine containing serotypes 1, 3, 6, 7, 14, 18, 19 and 23 (PCP-8). Group 1 received 25 microgram of each serotype at 3-6 months of age and again at 18-24 months. The antibody response after the second immunization was compared to a group of nine patients receiving a primary immunization at 18-24 months and to a group of ten age-matched controls receiving saline placebo. There were no significant differences in mean serum antibody levels between the two groups receiving the PCP-8. A fourth group of 44 infants between 6 and 21 months of age received either PCP-7 or PCP-8 and were followed for two years, at which time simultaneous injections of both vaccines were administered. Types 2, 3, 7, and 8 were most immunogenic but levels six months after immunization were approximately the same as for unimmunized controls with the exception of serotypes 3 and 7 which persisted for about two years. The class of antibody induced either by natural infection or by immunization was preferentially IgG and it was more often induced by the former. There were no significant differences between the serotypes of pneumococci isolated from nasopharyngeal cultures regardless of which vaccine was administered. Finally, the least immunogenic serotypes include 4, 6, 14, 19, and 23 and these are the only serotypes thus far associated with otitis media after immunization. The results suggest that PCP do not induce a lasting immune tolerance at the dose administered in this study; PCP are not very immunogenic in the young infant; PCP antibody tends to rise naturally; IgG antibody is preferentially induced; nasopharyngeal colonization is not altered by PCP immunization; and an association may exist between PCP immunogenicity and subsequent onset of otitis media.

Antigen-Antibody Reactions↗