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Biomedical subjects

S M Andrew

Publications and source records attributed to S M Andrew.

At least 19 recordsLinked to original sources
PubMed1999

An improved method for the structural profiling of keratan sulfates: analysis of keratan sulfates from brain and ovarian tumors.

A previously developed method for the structural fingerprinting of keratan sulfates (Brown et al., Glycobiology, 5, 311-317, 1995) has been adapted for use with oligosaccharides fluorescently labeled with 2-aminobenzoic acid following keratanase II digestion. The oligosaccharides are separated by high-pH anion-exchange chromatography on a Dionex AS4A-SC column. This methodology permits quantitative analysis of labeled oligosaccharides which can be detected at the sub-nanogram ( approximately 100 fmol) level. Satisfactory calibration of this method can be achieved using commercial keratan sulfate standards. Keratan sulfates from porcine brain phosphocan and human ovarian tumors have been examined using this methodology, and their structural features are discussed.

Acetylglucosaminidase↗
PubMed1997

Conversion of a traditional image archive into an image resource on compact disc.

The conversion of a traditional archive of pathology images was organised on 35 mm slides into a database of images stored on compact disc (CD-ROM), and textual descriptions were added to each image record. Students on a didactic pathology course found this resource useful as an aid to revision, despite relative computer illiteracy, and it is anticipated that students on a new problem based learning course, which incorporates experience with information technology, will benefit even more readily when they use the database as an educational resource. A text and image database on CD-ROM can be updated repeatedly, and the content manipulated to reflect the content and style of the courses it supports.

Archives↗
PubMed1997

Evaluation of selected antibiotic residue screening tests for milk from individual cows and examination of factors that affect the probability of false-positive outcomes.

Total composite milk samples from 131 cows in one herd were analyzed. Eight beta-lactam residue screening tests were evaluated for performance using milk from individual cows and factors that affect the rate of false-positive outcomes were determined. Cows were not treated with an antibiotic for at least 30 d prior to sampling. Tests evaluated were Delvotest P, Charm Cowside, Charm Farm, Penzyme, Valio T101, LacTek, CITE Probe, and Charm Bacillus stearothermophilus disk assay. Cows averaged 155 d of lactation. Milk production at the time of sampling ranged from 3.6 to 26.3 kg per milking per cow. The somatic cell count of milk averaged 243 x 10(3)/ml and ranged from 8.5 x 10(3)/ml to 3437 x 10(3)/ml. Total viable bacteria counts averaged 197.8 x 10(3)/ml. Total coliform counts ranged from 0 to 205/ml. Selectivity rates (rate of truly negative samples that were found to be negative by the assay) were greater than 90% for all tests except the CITE Probe test. Use of logistic regression showed that an increase in colony-forming units was associated with a decrease in the probability of a false-positive outcome for the CITE Probe test. Milk production, coliform counts, and parity each affected the probabilities of positive outcomes for different tests. Except for one test, selectivity rates of the beta-lactam residue screening tests for milk from individual cows was greater than 0.9.

Animals↗
PubMed1997

The Q-switched neodymium:YAG laser and tattoos: a microscopic analysis of laser-tattoo interactions.

The Nd:YAG laser effectively removes or lightens amateur and professional tattoos. The biomechanics of the removal of tattoo particles at the cellular level are incompletely understood. We examined biopsies obtained from 35 amateur and professional tattoos (including coloured tattoos), treated on three or more occasions with the Nd:YAG laser. Biopsies taken immediately after laser treatment showed vacuolation with complete clearance of tattoo particles in the most superficial layers of the dermis, as assessed by light and electron microscopy. We propose that the 'disappearance' of the tattoo particle arises from the formation of atomic species and gaseous products, which are rapidly dissolved in the extracellular fluid. Residual fragmented particles that are commonly found in the mid- and lower dermis are rephagocytosed. The interaction between the Nd:YAG laser and black tattoo particles at 1064 nm, and red tattoo particles at 532 nm, appears to be specific, as there was little evidence of thermal damage to adjacent cells or stromal collagen.

Humans↗
PubMed1995

Redirection of cellular cytotoxicity. A two-step approach using recombinant single-chain Fv molecules.

In this article the authors discuss an indirect system for redirecting cellular cytotoxicity, which utilizes a "universal" bispecific antibody to redirect T-cells to kill cells targeted with single-chain Fv (sFv) fusion proteins that carry a peptide tag recognized by the bispecific antibody. This approach has a number of theoretical advantages in the immunotherapy of cancer.

Antibodies, Bispecific↗
PubMed1995

Prediction of body composition of dairy cows at three physiological stages from deuterium oxide and urea dilution.

To develop equations for predicting body composition, mature Holstein cows (n = 21) were slaughtered at three physiological stages (-7, 63, and 269 d postpartum) after consecutive intravenous dosing with urea and D2O. Blood was sampled at 0 and 12 min after dosing with urea for determination of urea space and from 0 to 72 h after dosing with D2O. Empty body water and total body water were estimated by dilution kinetics for D2O using two- and one-compartment models, respectively. At slaughter, body components were ground, sampled, and freeze-dried for chemical analysis. Prediction of empty body water by urea space was not an improvement over the prediction by body weight alone. Prediction by D2O dilution explained 73 and 87% of the variation in empty and total body water, respectively. Estimated body protein, as determined from empty body water, predicted actual body protein with an error of 4.7 kg. Daily DMI explained 84% of the variation in the DM of the gastrointestinal tract contents (DM fill). Estimations of empty body fat (R2 = .85) and empty body energy (R2 = .89) from D2O dilution were capable of detecting significant differences in body fat (42.9 kg) and body energy (375 Mcal) across physiological stages and might be useful for prediction of body composition changes during the lactation cycle.

Animals↗
PubMed1994

Redirection of T cell-mediated cytotoxicity by a recombinant single-chain Fv molecule.

We have produced two single-chain Fv (sFv) proteins by bacterial periplasmic secretion, one sFv with specificity for the hapten DNP, and the other for the human transferrin receptor. After solubilization and refolding, we recovered several mg of active sFv per liter of bacterial culture. Each sFv bound to cells bearing the appropriate Ag and could be used to direct targeted cellular cytotoxicity. Targeting relied on a universal bispecific antibody designed to cross-link CD3 on the cytotoxic T cell with a peptide fused to the sFv carboxyl-terminus. The universal bispecific antibody was used in combination with the Ag-specific sFv to redirect human cytotoxic T cells to kill a variety of target cells. Such an approach has a number of advantages that may make it useful for the immunotherapy of cancer and other diseases.

Amino Acid Sequence↗
PubMed1994

Immunohistochemical appearance of corticosteroid contact hypersensitivity reactions.

We have studied, immunohistochemically, hypersensitivity reactions to corticosteroids and compared them with allergic contact dermatitis from nickel and appropriate controls. We could find no qualitative differences between nickel and corticosteroid contact reactions, providing further evidence that hypersensitivity to corticosteroids is an immunologically mediated reaction.

Administration, Topical↗
PubMed1994

Inflammatory cells in normal human fracture healing.

We studied inflammatory cells in specimens of callus taken from normally healing human fractures. Using immunohistochemistry, T-cells, B-cells, macrophages, HLA-DR expression and endothelial proliferation were assessed. Macrophages were present from an early stage but became less numerous later. T-cells were initially specifically recruited into the fracture site at the stage of granulation tissue, but subsequently excluded from areas of bone and cartilage formation. Inflammatory cells may control and coordinate fracture healing as has been proposed for soft tissue wound healing. The most likely mechanism for this is by the cytokines and growth factors which they are known to release.

Adult↗
PubMed1994

Direct analysis of body composition of dairy cows at three physiological stages.

Holstein cows were killed at three physiological stages, prepartum (-7 d, n = 10), early lactation (63 d, n = 7), and late lactation (269 d, n = 8), for determination of chemical composition and prediction of energy changes during lactation. Cows were weighed, slaughtered, and separated into five or six fractions, including carcass, gastrointestinal tract, mammary gland, uterus, and fetus (if present); the remainder was noncarcass. Live BW and weight of all empty body components except fat were unaffected by physiological stage. Empty body fat was reduced 42.3 kg for the early lactation cows compared with that of prepartum cows. Fat-free matter was similar across physiological stages; however, the water content of fat-free matter was greater for the prepartum and early lactation cows than for late lactation cows. In early lactation cows, the percentages of total protein were less in carcass and greater in gastrointestinal tissue than in prepartum and late lactation cows, but fat distribution was not affected. The energy values of 9.2 and 5.57 Mcal/kg for fat and protein in tissue, respectively, were determined by regression and used to apportion energy associated with fat, .925, and protein, .07, during lactation using data adjusted for ash. A maximum loss of 442 Mcal of tissue energy by approximately 77 DIM was determined by regression of empty body energy on DIM).

Animals↗
PubMed1993

An improved method for generating single-chain antibodies from hybridomas.

Cloning the correct VL kappa gene from hybridomas derived from MOPC-21 can be problematic because such cell lines variably express a transcript which is aberrantly rearranged at the VJ recombination site. Cellular levels of the aberrant transcript can exceed that of productive light chain RNA, so a large proportion of the VL gene-derived products obtained on PCR amplification of hybridoma cDNA may not encode a functional protein. We have developed a method in which antibody variable region genes are recovered from hybridoma cDNA using a unique set of V gene family-specific primers; the V region genes are then spliced by PCR, in the form 5'-VL-LINKER-VH-3' (where the linker encodes [GlyGlyGlyGlySer]3), and cloned into an expression vector under control of T7 RNA polymerase. Plasmid DNA is isolated from colonies, and the insert is expressed in an in vitro rabbit reticulocyte lysate-based coupled transcription/translation system, in a microtiter plate format. Since aberrantly rearranged VL kappa genes contain a translation termination codon at amino acid position 105, only constructs containing the correctly rearranged gene produce a protein of the predicted size. We demonstrate the method by producing the single-chain form of OKT9, a murine IgG1 which binds to the human transferrin receptor, and extend the results to show that the protein generated by the in vitro expression system retains the antigen binding properties of the parent antibody. Our method will be generally useful for screening single-chain antibodies for function prior to large scale production in vivo.

Amino Acid Sequence↗
PubMed1993

A bacterially expressed single-chain Fv construct from the 2B4 T-cell receptor.

A single-chain Fv construct of the 2B4 T-cell receptor has been made and expressed in Escherichia coli as bacterial inclusion bodies. After solubilization in 6 M guanidine hydrochloride and formation of mixed disulfides with glutathione, the protein was refolded by diluting out the denaturant and allowing intramolecular disulfide bridges to form by disulfide exchange. Approximately 65-100 mg of refolded protein was obtained from 1 liter of bacterial culture, an appreciable fraction of which was monomeric in nondenaturing solvents. This protein bound to three monoclonal antibodies specific for allotypic or idiotypic determinants on the native 2B4 variable region but did not bind several other anti-T-cell-receptor monoclonal antibodies that lacked such specificity. These experiments show that T-cell-receptor variable regions, like the V regions of antibodies, can form a well-behaved single-chain Fv molecule and provide large amounts of recombinant single-chain Fv T-cell receptor that can be used to study T-cell function.

Amino Acid Sequence↗
PubMed1993

Characterization of single-chain antibody (sFv)-toxin fusion proteins produced in vitro in rabbit reticulocyte lysate.

Chimeric proteins consisting of a fusion between binding-deficient mutants of diphtheria toxin (DT) or Pseudomonas exotoxin A (PE) and a single-chain antibody (E6 sFv) against the human transferrin receptor (TfnR) were expressed in a rabbit reticulocyte lysate system. Molecules utilizing PE40 (the carboxyl terminus 40 kDa of PE, lacking the binding domain) exhibited significant E6 sFv-mediated, cell type-specific cytotoxicity (IC50 1 x 10(-10) M) against a human erythroleukemia-derived cell line, K562. In contrast, a fusion protein between the same sFv and a DT mutant, DTM1 (containing two amino acid substitutions in the binding domain [S(508)F, S(525)F]) was not significantly cytotoxic, despite being enzymatically active. A tripartite protein in the form NH2-DTM1-E6 sFv-PE40-COOH exhibited cytotoxicity comparable to that of the PE40-sFv fusion (IC50 1 x 10(-10) M), suggesting that the deficit in activity of DTM1-sFv is not a function of misfolding of the sFv moiety or of a reduced ability to bind TfnR. In contrast to DTM1-E6 sFv, a fusion protein between a second DT mutant, CRM 107 [S(525)F], and the E6 sFv was specifically cytotoxic (IC50 1 x 10(-9) M), and toxicity could be blocked by addition of excess E6 antibody. The cell-free in vitro expression system we describe is rapid and may be used to express functional toxin-sFv fusion proteins. No protein refolding procedures are required, and the technique may be used to express proteins which, due to restrictions imposed on manipulation of toxin-encoding genes in Escherichia coli, could not be produced by more conventional methods.

ADP Ribose Transferases↗
PubMed1993

Demonstration of TGF-beta 1 mRNA by in situ hybridization in normal human fracture healing.

The role of transforming growth factor beta (TGF-beta) in fracture healing has previously been investigated in a rodent model, but not in human material. We investigated TGF-beta 1 gene expression in specimens of callus from normally healing human fractures, using in situ hybridization to a cDNA TGF-beta 1 probe and an autoradiographic disclosure system. TGF-beta 1 mRNA was present in areas of proliferation of mesenchymal tissue, bone, and cartilage. Levels of expression were lower in cells in the fracture hematoma and in differentiated (hypertrophic) chondrocytes. These results are compatible with those found in various animal models using immunohistochemistry and support the view that locally produced TGF-beta 1 is a regulator of fracture repair in humans from the early (mesenchymal proliferation) stage to the stage of remodeling of woven bone. They also indicate that, for TGF-beta 1, animal models accurately reflect human bone repair.

Adolescent↗
PubMed1993

Dermal dendrocytes in Dupuytren's disease: a link between the skin and pathogenesis?

The skin has previously been implicated in the process of Dupuytren's disease. The dermal dendrocyte is a factor XIIIa positive cell, which has been found in normal skin and some pathological conditions associated with fibrosis. In this study we examined the distribution of factor XIIIa positive cells in and around tissue from Dupuytren's disease. Immunohistochemistry was performed using a panel of antibodies for factor XIIIa, macrophages (CD68 and MAC387) and MHC II. Many factor XIIIa positive dendritic cells were present in and around Dupuytren's tissue; fewer CD68 and MHC II positive cells and very few MAC387 cells were seen. We propose that the factor XIIIa positive cells are dermal dendrocytes. This study may indicate an important link between the skin and pathogenesis of Dupuytren's disease.

Dendritic Cells↗
PubMed1993

Immunochemotherapy of human colon carcinoma xenografts in nude mice using combinations of idarubicin-monoclonal antibody conjugates.

Tumour cell heterogeneity is probably a principal cause of treatment failure and represents a formidable barrier for effective antibody-targeted chemotherapy. Idarubicin (Ida), a more potent and less cardiotoxic analogue of daunomycin, has been demonstrated to specifically target and eradicate homogeneous, cloned, murine tumour cell populations in vitro and in vivo when coupled to monoclonal antibodies (MoAb); however, the antitumour activity of Ida-MoAb conjugates against human tumour xenografts remains to be established. In this study, the value of cotargeting conjugates to different human tumour-associated antigens within a solid tumour has been assessed by comparing the effects of combinations of Ida-anti-colon carcinoma MoAb conjugates with any one Ida-anti-colon carcinoma MoAb conjugate used alone. Individual Ida-MoAb conjugates have previously been evaluated for their specific binding and cytotoxicity to one of two different human colon carcinoma xenografts (Colo 205 or LIM2210) in vitro, although their efficacy alone or in combination required assessment in vivo. Combinations of the most effective Ida-MoAb conjugates were demonstrated to enable a greater number of complete tumour regressions than the most efficacious Ida-MoAb conjugate administered alone in vivo; some combinations inhibited control tumour growth by up to 95%. This study suggests that Ida-MoAb conjugates can be effective against subcutaneous human tumours in nude mice, although it is unlikely that any single conjugate will eradicate all the tumour cells in a solid tumour, and the value of 'cocktails' of drug-MoAb conjugates against some xenografts (i.e. LIM2210) appears to be limited.

Animals↗