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S J Thorpe

Publications and source records attributed to S J Thorpe.

At least 19 recordsLinked to original sources
PubMed2006

An International Standard for specifying the minimum potency of anti-D blood-grouping reagents: evaluation of a candidate preparation in an international collaborative study.

BACKGROUND AND OBJECTIVES: The aim of this study was to evaluate a lyophilized monoclonal immunoglobulin M (IgM) anti-D preparation for use as an International Standard to specify a recommended minimum acceptable potency of anti-D blood-grouping reagents. MATERIALS AND METHODS: The candidate International Standard (99/836) for specifying the minimum potency of anti-D blood-grouping reagents was evaluated against a wide range of commercial anti-D blood-grouping reagents in an international collaborative study involving 20 laboratories in 13 countries. Laboratories titrated reconstituted 99/836, in parallel with as many commercial anti-D blood-grouping reagents as were available to them, in tube tests according to specified haemagglutination methodology for low-protein (e.g. monoclonal IgM) and high-protein (e.g. polyclonal) reagents. The ratios of the mean end-point titres of the reagents to that of 99/836 within each laboratory were calculated. RESULTS: The ratios of the mean titres of the low-protein reagents to the mean titre of 99/836 within a laboratory fell between 0.25 and 2 for 43 of the 45 low-protein anti-D reagents tested (i.e. the potencies of the low-protein reagents compared with 99/836 were between a 1:4 dilution of 99/836 to twice as potent as 99/836). The ratios of the mean titres of the high-protein reagents to the mean titre of 99/836 within a laboratory fell within 0.125 and 1 for eight out of the 10 high protein reagents tested. CONCLUSIONS: By international consensus, a 1:3 dilution of reconstituted 99/836 was deemed appropriate to define a recommended minimum acceptable potency of low-protein anti-D blood-grouping reagents. A 1:8 dilution of reconstituted 99/836 was deemed appropriate to define a recommended minimum acceptable potency of high-protein anti-D blood-grouping reagents. On the basis of the results presented here, 99/836 was established by the World Health Organization as the 1st International Standard for specifying the minimum potency of anti-D blood-grouping reagents, in tube tests.

Antibodies, Monoclonal↗
PubMed2006

International standards for minimum potency of anti-A and anti-B blood grouping reagents: evaluation of candidate preparations in an international collaborative study.

BACKGROUND AND OBJECTIVES: The aim of the study was to evaluate lyophilized monoclonal IgM anti-A and anti-B preparations for use as international standards (IS) to specify recommended minimum potencies of anti-A and anti-B blood grouping reagents in tube tests. MATERIALS AND METHODS: The candidate IS for minimum potency of anti-A and anti-B blood grouping reagents, codes 03/188 and 03/164, respectively, were evaluated against a wide range of commercial anti-A and anti-B blood grouping reagents in an international collaborative study involving 16 laboratories in nine countries. Laboratories titrated 03/188 and 03/164 in parallel with as many commercial anti-A and anti-B blood grouping reagents, respectively, as were available to them, in tube tests according to specified haemagglutination methodology. Three of these laboratories and a further laboratory also titrated 03/188 and 03/164 in parallel with currently available reference preparations for anti-A and anti-B. The ratios of the mean endpoint titres of the anti-A and anti-B reagents to those of 03/188 and 03/164, respectively, within each laboratory were calculated. RESULTS: The ratios of the mean titres of the anti-A reagents to the mean titre of 03/188 within a laboratory fell within 0.062 and 4, i.e. the potencies of the anti-A reagents were between a sixteenth to four times as strong as 03/188. The ratios of the mean titres of the anti-B reagents to the mean titre of 03/164 within a laboratory also fell within 0.062 and 4, with one outlier. CONCLUSIONS: By international consensus, a 1 in 8 dilution of the candidate IS for anti-A, 03/188, and a 1 in 4 dilution of the candidate IS for anti-B, 03/164, were considered appropriate to define the recommended minimum potencies of anti-A and anti-B blood grouping reagents, respectively, in tube tests. On the basis of these results, preparations 03/188 and 03/164 were established by the World Health Organization as International Standards for Minimum Potency of Anti-A and Anti-B Blood Grouping Reagents respectively, and by the US Food and Drug Administration Center for Biologics Evaluation and Research as Minimum Potency Reference Reagents.

ABO Blood-Group System↗
PubMed2005

International collaborative study to evaluate a candidate reference preparation to define an appropriate specified limit of anti-D in intravenous immunoglobulin products.

BACKGROUND AND OBJECTIVES: The aim of the study was to evaluate a lyophilized intravenous immunoglobulin (IVIG) preparation containing anti-D (02/228; nominal reciprocal titre of 8) for its suitability to define the maximum limit of anti-D in IVIG products when used in a proposed reference method of direct haemagglutination of papain-treated erythrocytes, in an international collaborative study. MATERIALS AND METHODS: Twenty laboratories tested 02/228 along with a negative control IVIG preparation and four IVIG samples containing different levels of anti-D. Nineteen laboratories performed direct haemagglutination methodology using papain-treated erythrocytes; five of these laboratories and one additional laboratory performed their in-house haemagglutination methodology (all indirect antiglobulin tests). RESULTS: The mode titre of 02/228, obtained by using the proposed reference method, was 8 (62.5% of tests). However, there was wide variation in haemagglutination titres between laboratories for three of the four samples. Correcting the titres of the samples relative to those of the proposed reference preparation reduced the interlaboratory variability and increased the frequency of the mode titres in three out of four samples. The indirect antiglobulin tests also showed wide interlaboratory variability and were less sensitive than the direct method in four laboratories. Eleven of the 14 laboratories that expressed an opinion considered that the level of anti-D in 02/228 was appropriate to define a specified limit. CONCLUSIONS: Our results demonstrate the necessity of using a reference preparation to define the maximum level of anti-D in IVIG products and ensure sufficient sensitivity in haemagglutination testing methodology. On the basis of these results, members of the European Pharmacopoeia Expert Group 6B recommended revision of the appropriate monograph to include this new specification and test. The Food and Drug Administration in the USA intends to adopt the same maximal specification defined by the reference preparation and to recommend the same test for the safety of IVIG products. Preparations 02/228 and 02/226 were also established by the World Health Organization as International Reference Reagents to standardize haemagglutination testing for anti-D in normal IVIG products.

Cooperative Behavior↗
PubMed2004

A calpain-like proteolytic activity produces the limited cleavage at the N-terminal regulatory domain of rabbit skeletal muscle AMP deaminase: evidence of a protective molecular mechanism.

On storage at 4 degrees C, rabbit skeletal muscle AMP deaminase undergoes limited proteolysis with the conversion of the native 85-kDa enzyme subunit to a 75-kDa core that is resistant to further proteolysis. Further studies have shown that limited proteolysis of AMP deaminase with trypsin, removing the 95-residue N-terminal fragment, converts the native enzyme to a species that exhibits hyperbolic kinetics even at low K+ concentration. The results of this report show that a 21-residue synthetic peptide, when incubated with the purified enzyme, is cleaved with a specificity identical to that reported for ubiquitous calpains. In addition, the cleavage of a specific fluorogenic peptide substrate by rabbit m-calpain is inhibited by a synthetic peptide that corresponds to residues 10-17 of rabbit skeletal muscle AMP deaminase; this peptide contains a sequence (K-E-L-D-D-A) that is present in the fourth subdomain A of rabbit calpastatin, suggesting that the N-terminus of AMP deaminase shares with calpastatin a regulatory sequence that might exert a protective role against the fragmentation-induced activation of AMP deaminase. These observations suggest that a calpain-like proteinase present in muscle removes from AMP deaminase a domain that holds the enzyme in an inactive conformation and which also contains a regulatory region that protects against unregulated proteolysis. We conclude that proteolysis of AMP deaminase is the basis of the large ammonia accumulation that occurs in skeletal muscle subjected to strong tetanic contraction or passing into rigor mortis.

AMP Deaminase↗
PubMed2004

Factors involved in the formation of attitudes towards those who are mentally ill.

BACKGROUND: This research was conducted to examine the prediction that accuracy of particular kinds of knowledge of mental illness would be related to particular attitudes towards people with mental illness. METHODS: An exploratory study examining attitudes to people suffering from mental illness is presented. A total of 169 participants, with a range of attitudes and knowledge, completed two questionnaires: attitudes were assessed using the Community Attitudes towards the Mentally Ill scale (Taylor and Dear 1981) which yields four attitude factors; knowledge was assessed using a questionnaire devised by Nunnally (1961), which yields ten knowledge factors. RESULTS: Multiple regression analyses provided some support for the hypothesis that some areas of knowledge (particularly knowledge concerning guidance and support, and knowledge concerning the role of avoidance of morbid thoughts in mental health) are predictive of specific attitudes, but much of the variance remained unexplained by the predictive knowledge variables. Post hoc analyses revealed that those people who had had personal experience of people with mental illness were generally more positive in their attitudes towards them. CONCLUSIONS: Selected knowledge factors only accounted for a modest amount of the variance in attitude factor scores. Affective (e. g. fear, revulsion, anxiety) information may explain a greater percentage of variance in attitude factor score. It is suggested that this should be considered in future research and the limitations of the present study are discussed.

Adolescent↗
PubMed2004

Collaborative study for establishment of a global standard for the potency assay of human anti-D immunoglobulin.

An international collaborative study aimed at establishing a global standard for the potency assay of anti-D immunoglobulin was started in 2002. 25 laboratories participated in this study run under the common aegis of the World Health Organization, the United States Food and Drug Administration (US-FDA) and the European Directorate for the Quality of Medicines (EDQM). The potencies of three candidate materials and the US-FDA standard (lot 3) included for comparison were evaluated using AutoAnalyzer, competitive enzyme-linked immunoassay (competitive EIA), flow cytometric methods or own "in-house" methods. Critical reagent, standardised procedures and standardised assay design were provided for either method, where appropriate. Central statistical evaluation of the potency data submitted by the participants was performed using a parallel line model. Agreement between laboratories and assay methods for all samples was observed. Intra-laboratory variability was lowest for laboratories performing flow cytometry and highest for laboratories that performed their in-house methods. Inter-laboratory variability was acceptable for all samples when assayed by AutoAnalyzer, competitive (EIA) and flow cytometric methods. It was concluded that sample A is most suitable to serve as a global standard and that sample C could serve as a reserve European Pharmacopoeia (Ph. Eur.) Biological Reference Preparation (BRP) batch provided that suitable stability is demonstrated. Sample A was adopted by the Ph. Eur. Commission at its 115th session (March 2003) as the first Ph. Eur. BRP (available from the EDQM: catalog number Y0000219) with the assigned potency of 285 IU/ampoule.

Chromatography, High Pressure Liquid↗
PubMed2003

The relationship between disgust sensitivity, anxiety and obsessions.

Three hundred participants, including volunteers from an obsessional support group, filled in questionnaires relating to disgust sensitivity, health anxiety, anxiety, fear of death, fear of contamination and obsessionality as part of an investigation into the involvement of disgust sensitivity in types of obsessions. Overall, the data supported the hypothesis that a relationship does exist between disgust sensitivity and the targeted variables. A significant predictive relationship was found between disgust sensitivity and total scores on the obsessive compulsive inventory (OCI; Psychological Assessment 10 (1998) 206) for both frequency and distress of symptomatology. Disgust sensitivity scores were significantly related to health anxiety scores and general anxiety scores and to all the obsessional subscales, with the exception of hoarding. Additionally, multiple regression analyses revealed that disgust sensitivity may be more specifically related to washing compulsions: frequency of washing behaviour was best predicted by disgust sensitivity scores. Washing distress scores were best predicted by health anxiety scores, though disgust sensitivity entered in the second model. It is suggested that further research on the relationship between disgust sensitivity and obsessionality could be helpful in refining the theoretical understanding of obsessions.

Adult↗
PubMed2003

Neutralizing increases discomfort associated with obsessional thoughts: an experimental study with obsessional patients.

Cognitive-behavioral theories suggest that the development of neutralizing is crucial in the development and persistence of obsessional problems (OCD). Twenty-nine patients with a Diagnostic and Statistical Manual of Mental Disorders (4th ed., American Psychiatric Association, 1994) diagnosis of OCD were randomly allocated to 2 conditions. Both listened to repeated recorded presentations of their intrusive thoughts and either neutralized (experimental group) or distracted themselves (control). Discomfort was rated during this 1st phase and then during a 2nd phase without neutralizing or distraction. The experimental group showed a similar level of discomfort in the 1st phase, which significantly reduced during the period compared with controls. The experimental group experienced significantly more discomfort during the 2nd phase, and significantly stronger urges to neutralize and distract at the end of this phase than controls.

Adult↗
PubMed2003

Clonal analysis of a human antimouse antibody (HAMA) response.

Circulating human antimouse antibodies (HAMAs) directed to mouse immunoglobulin G (IgG) are clinically significant, compromising mouse antibody therapy and imaging, and interfering in immunological assays. To investigate the HAMA response, 20 stable cell lines secreting human monoclonal antibodies reactive with mouse IgG were established from a donor with a history of exposure to mice. Their subclass and domain specificities were established by solid-phase binding, indirect haemagglutination assays and immunoblotting, using Igs of known subclass and Ig fragments. The heavy-chain variable region gene usage was determined for 12 HAMAs. Eight HAMAs were IgM, 11 HAMAs were IgG4 and one HAMA was IgG1, indicating an IgG4-dominated response. All of the IgG HAMAs reacted with epitopes present on the Fc portion; one was subclass-specific, nine were subclass-restricted and two were pan-IgG-reactive. Measurement of their affinities gave dissociation constants typically in the nanomolar range. Seven and five HAMAs were derived from variable heavy-chain 3 (VH3) and VH1 gene segments, respectively. The IgG HAMAs used different VH segments to the IgM HAMAs. JH regions were coded by JH4 in eight HAMAs. DH segment usage appeared to be restricted in the IgM HAMAs. Two IgG HAMAs were clonally related. These monoclonal HAMAs are potentially useful as reagents for detecting mouse IgG and as reference reagents for the investigation of the HAMA response in patients undergoing mouse monoclonal antibody therapy and for the investigation of the influence of HAMAs on immunodiagnostic tests.

Amino Acid Sequence↗
PubMed2003

Competitive enzyme-linked immunoassay of monoclonal immunoglobulin G anti-D preparations.

The development of monoclonal immunoglobulin G (IgG) anti-D for prophylaxis necessitates estimation of their potency in terms of their red cell-binding ability, but it is unclear which quantification methodology is most suitable for this. The aim of this study was to assess 50 monoclonal anti-D from the 4th International Workshop for quantitative and qualitative binding to red cells in a competitive enzyme-linked immunoassay (EIA) in which varying amounts of monoclonal antibodies (MoAbs) and a constant amount of biotinylated monoclonal anti-D (BRAD-5) compete for red cell binding. The potencies of the MoAb were estimated against the International Reference Preparation (IRP) for anti-D Ig. Two MoAbs as supplied were insufficiently inhibitory of biotinylated BRAD-5 binding to be quantified; the potencies of the remainder ranged from 4 to 343 IU mL-1 and included 11 MoAbs showing dose-responses that were nonparallel to those of other MoAb and the IRP. Estimation of the concentration of antibody in the supernatants by radial immunodiffusion ranged from 0.5 to 61 micro g mL-1, giving specific activities of <1-24 IU micro g-1. The results show that competitive EIA is suitable for quantitating most monoclonal anti-D for development and quality control purposes, regardless of their D epitope reactivity.

Antibodies, Monoclonal↗
PubMed2003

Batches of intravenous immunoglobulin associated with adverse reactions in recipients contain atypically high anti-Rh D activity.

BACKGROUND AND OBJECTIVES: The presence of anti-Rh D in intravenous immunoglobulin (IVIG) products has been claimed to be associated with adverse reactions in recipients. There is currently no regulatory specification to control the level of anti-D in IVIG products and it is unclear what this should be. Two reports of haemolysis occurring in recipients of IVIG manufactured from US plasma provided a rare opportunity to investigate whether high anti-D levels could have induced the haemolysis. MATERIALS AND METHODS: We developed a direct microtitre plate haemagglutination method suitable for screening IVIG products and starting plasma pools for haemagglutinating activity. RESULTS: Of 101 batches of IVIG tested, six were found to contain specific anti-D. Four of these batches had anti-D titres ranging from 64 to 256 (including the two batches each associated with a report of haemolysis) and could be linked, in each case, to a starting plasma pool also positive for anti-D. CONCLUSIONS: Our results show that IVIG products can contain appreciable anti-D levels. To avoid potential problems in recipients, we propose an anti-D titre of 8 as the maximum permissible limit of anti-D in IVIG products for batch acceptance and release. The availability of a reference preparation is essential for control of this proposed requirement.

Blood Cells↗
PubMed2003

A global standard for anti-D immunoglobulin: international collaborative study to evaluate a candidate preparation.

BACKGROUND AND OBJECTIVES: The aim of the study was to evaluate a lyophilized anti-D immunoglobulin preparation to serve as a global standard for potency assays of anti-D immunoglobulin products. MATERIALS AND METHODS: The candidate global standard, 01/572, was calibrated against the World Health Organization (WHO) International Reference Preparation (IRP) for anti-D immunoglobulin, human (68/419), along with two reserve candidate reference preparations, in an international collaborative study involving 25 laboratories in 15 countries. The United States Food and Drug Administration (US-FDA) Center for Biologics Evaluation and Research (CBER) Standard for anti-D immunoglobulin, Lot 3, was included for comparison. Most laboratories (20/25) performed AutoAnalyser methodology, competitive enzyme-linked immunoassay (EIA) and/or flow cytometry. RESULTS: The overall mean potency of the candidate global standard, 01/572, was 284.5 international units (IU)/ampoule, with an interlaboratory variability, expressed as a percentage geometric coefficient of variation (% gcv), of 9.7. The mean potency of the US Standard was 859.4 IU/ml with an interlaboratory variability of 9.5% gcv, excluding an outlier. The mean potencies of the reserve preparations per ampoule/vial were 110.6 IU and 106.7 IU when calibrated against the IRP, and 112.2 IU and 106.6 IU when calibrated against 01/572, respectively, with interlaboratory % gcv values of 9.6-18.3 (excluding outliers). CONCLUSIONS: Preparation 01/572 proved more suitable for use as a global standard than the reserve candidate preparations and was established, with an assigned potency of 285 IU/ampoule, by the WHO as the 2nd International Standard for anti-D immunoglobulin; by FDA-CBER as the Standard for anti-D immunoglobulin, Lot 4; and by the European Directorate for the Quality of Medicines (EDQM) as the 1st Biological Reference Preparation for anti-D immunoglobulin.

Australia↗
PubMed2002

International collaborative study to evaluate methods for quantification of anti-D in immunoglobulin preparations.

BACKGROUND AND OBJECTIVES: The disadvantages of autoanalyser methodology for anti-D potency estimation have prompted the search for an alternative reference method. The aim of this study was to carry out a direct comparison of autoanalyser methodology, competitive enzyme-linked immunoassay (EIA) and flow cytometry. MATERIALS AND METHODS: The anti-D potencies of nine immunoglobulin preparations were estimated against the World Health Organization (WHO) International Reference Preparation for anti-D immunoglobulin, using the three different assay methods described above, in an international collaborative study involving 18 laboratories. RESULTS: No significant differences in potency estimates for five of nine samples were identified using the three different methods. For six of nine samples the mean potency estimates obtained using competitive EIA lay between those of the autoanalyser and flow cytometry. Flow cytometry gave the lowest estimates for seven of nine samples. The range of intralaboratory variability, expressed as percentage geometric coefficients of variation (% gcv) and excluding extreme values, was as follows: autoanalyser (eight laboratories), 0.4-17.0; competitive EIA (12 laboratories), 2.6-34.8; and flow cytometry (eight laboratories), 2.9-30.0. The range of interlaboratory variability (expressed as % gcv) was: autoanalyser (eight laboratories, eight samples), 10.8-17.6; competitive EIA (12 laboratories, nine samples), 10.3-17.3; flow cytometry (nine laboratories, eight samples), 6.2-16.1. CONCLUSIONS: The study did not show clear superiority of one method over the others. Both competitive EIA and flow cytometry are acceptable alternatives to autoanalyser methodology for polyclonal anti-D potency estimation. For monoclonal anti-D, further work is necessary to determine the most appropriate method for potency testing.

Agglutination Tests↗
PubMed2001

The time course of visual processing: from early perception to decision-making.

Experiments investigating the mechanisms involved in visual processing often fail to separate low-level encoding mechanisms from higher-level behaviorally relevant ones. Using an alternating dual-task event-related potential (ERP) experimental paradigm (animals or vehicles categorization) where targets of one task are intermixed among distractors of the other, we show that visual categorization of a natural scene involves different mechanisms with different time courses: a perceptual, task-independent mechanism, followed by a task-related, category-independent process. Although average ERP responses reflect the visual category of the stimulus shortly after visual processing has begun (e.g. 75-80 msec), this difference is not correlated with the subject's behavior until 150 msec poststimulus.

Adult↗
PubMed2001

Enhancing the catalytic repertoire of nucleic acids. II. Simultaneous incorporation of amino and imidazolyl functionalities by two modified triphosphates during PCR.

The incorporation of potentially catalytic groups into DNA is of interest for the in vitro selection of novel deoxyribozymes. We have devised synthetic routes to a series of three C7 modified 7-deaza-dATP derivatives with pendant aminopropyl, Z-aminopropenyl and aminopropynyl side chains. These modified triphosphates have been tested as substrates for Taq polymerase during PCR. All the modifications are tolerated by this enzyme, with the aminopropynyl side chain giving the best result. Most protein enzymes have more than one type of catalytic group located in their active site. By using C5-imidazolyl-modified dUTPs together with 3-(aminopropynyl)-7-deaza-dATP in place of the natural nucleotides dTTP and dATP, we have demonstrated the simultaneous incorporation of both amino and imidazolyl moieties into a DNA molecule during PCR. The PCR product containing the four natural bases was fully digested by XbaI, while PCR products containing the modified 7-deaza-dATP analogues were not cleaved. Direct evidence for the simultaneous incorporation during PCR of an imidazole-modified dUTP and an amino-modified 7-deaza-dATP has been obtained using mass spectrometry.

Binding Sites↗
PubMed2001

Enhancing the catalytic repertoire of nucleic acids: a systematic study of linker length and rigidity.

The incorporation of potentially catalytic groups in DNA is of interest for the in vitro selection of novel deoxyribozymes. A series of 10 C5-modified analogues of 2'-deoxyuridine triphosphate have been synthesised that possess side chains of differing flexibility and bearing a primary amino or imidazole functionality. For each series of nucleotide analogues differing degrees of flexibility of the C5 side chain was achieved through the use of alkynyl, alkenyl and alkyl moieties. The imidazole function was conjugated to these C5-amino-modified nucleotides using either imidazole 4-acetic acid or imidazole 4-acrylic acid (urocanic acid). The substrate properties of the nucleotides (fully replacing dTTP) with TAQ polymerase during PCR have been investigated in order to evaluate their potential applications for in vitro selection experiments. 5-(3-Aminopropynyl)dUTP and 5-(E-3-aminopropenyl)dUTP and their imidazole 4-acetic acid- and urocanic acid-modified conjugates were found to be substrates. In contrast, C5-amino-modified dUTPs with alkane or Z-alkene linkers and their corresponding conjugates were not substrates. The incorporation of these analogues during PCR has been confirmed by inhibition of restriction enzyme digestion using XBAI and by mass spectrometry of the PCR products.

Catalysis↗
PubMed2001

Enhanced efficacy of anti-D immunoglobulin for treating ITP is not explained by higher immunoglobulin polymer content.

Several reports have suggested that low dose anti-D immunoglobulin is superior to high dose immunoglobulin for treatment of idiopathic thrombocytopenia purpura (ITP). However, some findings suggest that it is not the anti-D activity per se that is responsible for efficacy for treatment of ITP with anti-D immunoglobulin. Amongst alternative explanations for the mechanism of action is a relatively higher immunoglobulin polymer content of anti-D compared to other immunoglobulin products, which is more efficient in causing reticuloendothelial Fc receptor blockade. In order to investigate this we have evaluated the polymeric IgG content of anti-D and other immunoglobulin products. Different products showed considerable variation in immunoglobulin polymer content. There was no clear correlation between aggregate or dimer content and product type and anti-D as a class of product did not contain higher amounts of either dimer or aggregate compared to other products. Some manufacturers' products increased in polymer content on storage, but others did not show this effect. Therefore, higher immunoglobulin dimer and/or aggregate content cannot explain the increased efficacy of anti-D immunoglobulin for treatment of ITP. The role of polymeric Ig in efficacy for treatment of ITP is unclear.

Biopolymers↗