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Biomedical subjects

A R Brown

Publications and source records attributed to A R Brown.

At least 55 records · Page 3Linked to original sources
PubMed1992

Stereoselective high-performance liquid chromatographic assay with fluorometric detection for the isomers of mivacurium in human plasma.

A high-performance liquid chromatographic assay with fluorometric detection was developed for the analysis of the stereoisomers of mivacurium, a new short-acting neuromuscular blocker, in plasma. The isomers were isolated from plasma by solid-phase extraction with C18 and anion-exchange cartridges. The extracts were chromatographed on a LiChrosphere 60 RP Select B column (125 mm x 4.6 mm I.D.) using a mobile phase of acetonitrile-water (4:6, v/v) containing 0.005 M octanesulfonic acid. The fluorescence excitation and emission wavelengths were 202 and 320 nm, respectively. The accuracy and precision of the assay, expressed as the percentage deviation of measured values from true values and the percentage coefficient of variation, respectively, were less than or equal to 10% at all concentrations except for the percentage coefficient of variation at the lower limit of quantitation (5 ng/ml). The assay has been successfully used for the analysis of plasma samples from a pharmacokinetic study in human volunteers.

Chromatography, High Pressure Liquid↗
PubMed1992

In situ detection of autoanti-idiotype antibody-forming cells induced by influenza virus infection.

In situ immunocytochemical-staining methods combined with computer-aided image analysis were employed to examine autoanti-idiotype antibody-forming cell expansion in vivo. Autoanti-idiotype antibody-forming cells were demonstrated in the spleens of C57BL/6J (B6) strain mice intranasally infected with the influenza virus A/Hong Kong/168/(H3N2)[R] X-31. Autoanti-idiotype B cells were detected and elevated in spleen tissues after secondary influenza infections compared to normal B6 mice, and were specific for a dominant idiotype antibody called PY206 reactive with the influenza virus hemagglutinin. Influenza virus-infected BALB/c mice, which make little PY206 Id, did not have increased autoanti-Id against PY206 compared to normal mice. Results provide evidence for an idiotype network-mediated stimulation of autoanti-idiotype B cell production during influenza virus infection.

Animals↗
PubMed1992

Immunological functions of splenic B-lymphocytes.

The spleen is a key lymphoid organ for generating B-lymphocyte (B cell)-mediated humoral immunity. This review examines the key features and functions of splenic B cells. Splenic B cell subsets, including virgin, memory, and CD5+ B cells, are characterized by their phenotypic markers and functions. Structural aspects of the spleen, including red pulp, follicles, periarterial lymphoid sheaths (PALS), and germinal centers, are related to the B cells localized in these areas. The migratory behavior of B cells in these splenic compartments is considered in the context of antigen exposure, adhesion molecules, and migratory stimuli. Antigen-specific B cells in the spleen are examined with an emphasis on their in situ characteristics assessed by immunocytochemical methods. This includes the detection of antigen-binding antibody-forming cells (AFC), idiotype-producing AFC, and the application of computer-aided imaging to analyze these cells. Interactions between splenic B cells and other cell types, such as T-lymphocytes (T cells) and antigen-processing/presenting cells like macrophages and dendritic cells, are briefly examined. Cytokines that influence splenic B cells are reviewed. Interactions between B cells and the neuroendocrine system are discussed briefly. The functions of splenic B cells are considered. Antibody production, cytokine synthesis, and potential immunoregulatory activities are considered.

Antibody Formation↗
PubMed1991

A dominant idiotype in the antibody response against the influenza virus hemagglutinin. Serum and in situ analyses.

PY206 is an Id associated with a BALB/c murine mAb described as being specific for the influenza A virus hemagglutinin. However, production of this Id by BALB/c mice immunized with influenza is low. This report shows that the PY206 Id is a dominant component of the anti-influenza antibody response in C57BL/6J strain mice infected intranasally with the influenza A/Hong Kong/168/(H3N2)[R] X-31 virus. High PY206 Id expression was linked to the IgHb Ig allotype locus. PY206 Id+ antibody-forming cells were identified in situ in cryostat sections of lymphoid tissues and idiotypic heterogeneity was identified among PY206+ B cells. Uninfected adult C57BL/6J mice had PY206 Id in their serum that lacked influenza binding specificity. In situ analysis of prenatal and neonatal spleen of uninfected C57BL/6J mice showed that the expansion of PY206 Id+ B cells occurred early in development. PY206+ cells were demonstrated in the lungs of influenza-infected mice but not in normal mice, establishing the capability to study this B cell population in the lung. This model offers the opportunity to manipulate the anti-influenza A virus hemagglutinin B cell response and to study the proliferation and migration of influenza-specific B cells in their native tissue environments.

Administration, Intranasal↗
PubMed1990

Tumor necrosis factor-alpha analyzed within individual macrophages by combined immunocytochemistry and computer-aided image analysis.

Immunocytochemical staining procedures were combined with computer-aided image analysis to quantitate the relative intracellular production of tumor necrosis factor-alpha (TNF) within individual macrophages. Optimal conditions for time and methods for the activation of TNF production, fixation of cells for optimal immunocytochemical staining, and image analysis methods were determined. Thioglycolate elicited peritoneal macrophages were readily activated to significantly increased levels of intracellular TNF, as early as 1 hr after activation with lypopolysaccharide (LPS) + interferon-gamma: maximum intracellular TNF was evident after 2-3 hr. Both LPS and interferon-gamma was necessary to increase intracellular TNF. Normal alveolar macrophages also readily produced increased intracellular TNF, but normal peritoneal and splenic macrophages were poorly activated to TNF production. Acid stripping of receptor bound TNF allowed discrimination between intracellular TNF/integral membrane TNF, and TNF-receptor-bound TNF. Results stress the importance of studying these TNF forms early after activation. Applications for TNF quantitation by these means are discussed.

Animals↗
PubMed1990

Prevalence of maternal HIV infection based on anonymous testing of neonates, Sydney 1989.

The presence of antibody to human immunodeficiency virus (HIV) in post-partum women may be inferred by screening the blood of their newborn babies, since maternal IgG antibodies freely cross the placenta. We tested a sample of 10,217 newborns from 10 hospitals covering three areas in Sydney and other metropolitan centres in New South Wales from April to July, 1989. None of the specimens gave a positive test for antibody to HIV. Thus, the prevalence of HIV positive serology in this sample of newborns was found to be zero. It was estimated that the seroprevalence of antibody to HIV among all neonates in the study area was between zero and 0.045% (99% confidence interval). Because newborns are an accessible group for the study of HIV, and can act as surrogates for their mothers, anonymous testing of this sentinel group will remove some of the limitations generalizing the information in the present database of HIV infection in Australia. This study provides baseline data and suggests that there is not a widespread epidemic of HIV infection among heterosexual persons in Australia at the present time and that routine antenatal testing of women for antibody to HIV may not be cost-effective. However, it will be important to repeat this study at regular intervals to detect any increase in HIV seroprevalence.

Adult↗
PubMed1990

Combined immunocytochemical staining and image analysis for the study of lymphocyte specificity and function in situ.

Immunocytochemical staining methods were combined with computer-aided image analysis to quantitate relative intracytoplasmic and cell membrane products within single lymphoid cells. Lymphoid cells fixed to microscope slides were immunocytochemically stained and the densities of immunocytochemical stains for individual lymphoid cells were determined on video-camera-captured microscopic images to quantitate synthesized and fully assembled antibody molecules detected by an anti-idiotypic antibody. Similarly, expression of a lymphoid cell membrane molecule, Ly-1, was comparatively quantitated in individual lymphoid cells. Densitometric analysis of tissue sections was utilized to measure tissue areas and quantitate cell nuclei in tissues. Two-color immunocytochemical staining and sequential image analyzer-determined locations of individual cells in tissues allowed unambiguous identification of doubly stained cells in their native tissue environment. This analysis was applied to the in situ identification of antibody forming cells producing the CRIc idiotype of the BALB/c anti-arsonate response and a determination of the mu chain usage by those individual cells. Applications of combined immunocytochemistry and computerized image analysis to studies of in vivo immunologic functions are discussed.

Animals↗
PubMed1989

In situ analysis of antibody-forming cells from the BALB/c CRIc idiotype family: idiotopic heterogeneity among clustered cells.

Immunocytochemical staining methods were applied toward defining the in situ expansion of the BALB/c mouse antibody-forming cells (AFC) that express the CRIc idiotype (id) family associated with the antibody response against the p-azophenylarsonate (Ar) hapten. CRIc+ AFC dominated the early primary anti-Ar response but represented a decreased fraction of the anti-Ar AFC during secondary responses. Two subsets of the CRIc family, CRIc1 and CRIc2, differed in their relative expression, with CRIc1 more pronounced during primary responses and CRIc2 better expanded in secondary responses. Idiotopic differences among AFC in very close proximity suggested convergent migration among B cells having similar but not identical V regions or possibly V region mutations among clonally derived cells. The approach also allowed in situ idiotypic analysis of polyclonally activated B AFC.

Adjuvants, Immunologic↗
PubMed1988

Detection of antibody, idiotype, and anti-idiotype forming cells by in situ immunocytochemical staining.

Methods are described for the immunocytochemical staining of cryostat sections of lymphoid tissue with enzyme conjugates of antigen, idiotype (Id) and anti-idiotype. Results established this as a useful approach, for simultaneously detecting Id and anti-Id antibody forming cells (AFC) in situ. As a model, the 5AF6 Id family associated with the BALB/c mouse antibody response against the p-azophenyl-arsonate (Ar) epitope was examined by two-color immunocytochemical staining, allowing the simultaneous detection of both Id+ and Id- anti-Ar AFC. Spleens from mice secondarily immunized with Ar antigen but not normal mice contained anti-Id AFC stained with the 5AF6 Id but not with another immunoglobulin of the same isotype. A sequential staining method was developed which allowed the detection of both Id and anti-Id AFC in the same tissue, thus providing a means of examining Id and anti-Id antibody networks in intact lymphoid tissues.

Animals↗
PubMed1988

Newborn screening for cystic fibrosis using an enzyme linked immunoabsorbent assay (ELISA) technique.

Neonatal screening for cystic fibrosis (CF) has been conducted using conventional radioimmunoassay. An alternative immunoassay approach has been developed and applied to screening 35,550 newborns. Seventeen confirmed CF infants were detected by both assays. This monoclonal antibody-based enzyme immunoassay using microtitre plate ELISA technology has proved effective in case finding and offers a number of advantages. A reduced labour component, ease of handling, use of nonradioactive reagents, long reagent shelf-life, greater specificity and a reduction in potential sample handling and transposition errors combine to make this technology appropriate for a large volume neonatal screening laboratory.

Antibodies, Monoclonal↗
PubMed1987

Two BALB/c anti-arsonate idiotype families: two heavy chain variable regions (Vh) shared with anti-DNP antibodies are used by one family while a Vh similar to anti-GAT antibodies is used by the other.

Amino terminal amino acid sequences were obtained for both the heavy (H) and light (L) chains of seven BALB/c anti-arsonate (Ar) monoclonal antibodies representing the 5AF6 and 3C6 idiotype (id) families described in this strain. 5AF6 family H chains showed strong homology to the germ-line gene sequence for the A strain 36-60 family. However, four to five identical H chain sequence differences for two of these antibodies (5AF6 and 95B5), as well as two previously reported related sequences (92D5, 94B10), suggested they were encoded by a different Vh. The 36-60 family Vh genes have been shown to be identical to the Vh gene of the anti-DNP binding myeloma M460 [Dzierzak et al., J. Immun. 136, 1864-1870 (1986)]. H chains amino acid sequences derived from an id-460+ anti-DNP hybridoma and a germ-line gene differing from the 30-60-like Vh sequence [Dzierzak et al., J. Immun. 136, 1864-1870 (1986)] were found to be virtually identical to the 95B5 and 5AF6 Vh sequences. This suggests that the same two related H chains making up two subsets of the 5AF6 anti-Ar id family are also both used in two subsets of the id-460 anti-DNP response. 5AF6 family L chains were highly homologous to the other Vk2 L chains of the 36-60 family. 3C6 family H chains can be placed in the Vh l group, are unrelated to the described anti-Ar H chain families, and have been placed in a new anti-Ar Vh family, Ars-E. The 3C6 H is similar, however, to a Vh used by a BALB/c anti-GAT idiotype family of antibodies. 3C6 L chains were of the murine kappa chain group, Vk8 and most resembled an L chain from an A strain monoclonal anti-Ar having no defined idiotype.

Amino Acid Sequence↗
PubMed1986

Regulation of the BALB/c anti-p-azophenylarsonate antibody response by monoclonal anti-idiotype. I. Anti-idiotope fine specificity.

Five monoclonal anti-idiotype antibodies were prepared against the IgG1 monoclonal antibody, 5AF6, the prototype molecule representing the BALB/c 5AF6 idiotype family of antibodies specific for the p-azophenylarsonate (Ar) hapten. Three were of BALB/c origin and two were derived from allotype congenic strain CB.20. All five anti-idiotopes (id) reacted with the 5AF6 immunogen but not with four other BALB/c anti-Ar sharing other id with 5AF6. Four of the five showed some reactivity with three monoclonal anti-Ar derived from A strain mice that represent a minor component of the anti-Ar from that strain. Reactivity patterns of these anti-id indicated that all five reacted with different id on the 5AF6 molecule, yet all five were sufficiently close to the Ar-binding site for their binding to be blocked by the Ar hapten alone. Furthermore, all five anti-id could compete with each other for binding to 5AF6, indicating that the five id detected by these anti-id were in close proximity. Four of the five anti-id reacted with id produced by conformations requiring both the appropriate heavy and light chains. The fifth anti-id reacted with a heavy chain id stabilized by the presence of any light chain. The implications of such a diverse anti-id response against a single antibody molecule on anti-id network interactions are discussed.

Animals↗
PubMed1986

Regulation of the BALB/c anti-p-azophenylarsonate antibody response by monoclonal anti-idiotype. II. Idiotope expression in serum and its regulation.

Five murine monoclonal antibody (mAb) anti-idiotypes (id), shown in the accompanying report by binding studies to be reactive with five different id on a single member of the 5AF6 family of BALB/c antibodies against the p-azophenylarsonate (Ar) hapten, were used to examine the distribution of their recognized id among anti-Ar in BALB/c and other mouse strains immunized with keyhole limpet hemocyanin-Ar (KLH-Ar). Differences in id expression in BALB/c and other strains substantiate that all five monoclonal anti-id reacted with different id. This suggests that the anti-id repertoire for a single antibody molecule may be extensive. Two of the anti-id reacted with id that were found in virtually all KLH-Ar immunized BALB/c mice, but constituted only a subset (approximately 33%) of the antibodies representing the 5AF6 family. The other three anti-id reacted with id infrequently expressed among BALB/c anti-Ar. Other mouse strains producing 5AF6 family anti-Ar antibodies also produced antibodies recognized by mAb 2CB8 and 6BA1; however, the three id infrequently expressed in BALB/c mice were produced in higher quantities and in a greater percent of mice. Monoclonal anti-id were capable of suppressing a portion but not all of the 5AF6 family of anti-Ar antibodies. Four of the five anti-id suppressed a greater fraction of the 5AF6 family than that id represented in a normal immune response, suggesting that suppression was mediated via an id other than that recognized by these monoclonal anti-id. Overall, the results indicate that an extensive repertoire of anti-id can be produced against a single id antibody, but suppression induced by treatment with these anti-id in this model is presumably mediated via another as yet unidentified id determinant(s).

Animals↗